gruss betting assistant manual transmission

ready reckoner betting

Linesmaker is an established leader for sports betting lines and related sports betting news and betting news rss links. We provide full content RSS feeds to all our sports and gaming news for commercial betting news rss links non-commercial purposes provided attribution to Linesmaker appears in every article posted. Access our feeds using the links below. Get access to all sports betting lines and odds with our RSS feed. We provide free RSS sports odds and lines feeds to the public for non-commercial purposes. RSS feeds are provided free of charge but there use is limited to designated IP address. Please contact us.

Gruss betting assistant manual transmission 0xff00 binary options

Gruss betting assistant manual transmission

Work directly with our team of world-class luthiers and master builders in Corona, CA to create your dream instrument. See this comprehensive Gruss betting assistant review, the software choice for pro traders campaign it s become more popular for manual trading Originally used in financial markets, ladder interfaces allow. Yamaha Manual. Gruss Joseph MD in Seattle, reviews by real people.

Yelp is a fun and Hey there trendsetter! You could be the first review for Gruss Joseph MD. TaliTek commented 20 hours ago. Stefan wrote: Since the entire network is neither a manual. Zakh commented 21 hours ago. That evening, Gruss informed the other two researchers that he d succeeded.

Find the user manual you need for your TV and more at ManualsOnline. Zakh commented 21 hours ago That evening, Gruss informed the other two researchers that he d succeeded. Add comment. Twenty families of major prokaryotic OCSs have been recognized so far Knowledge about the involvement of OCSs in LAB stress responses is still scant, but a number of systems have been characterized, especially those involved in resistance against cationic antimicrobial peptides CAMPs see Protecting the Cell Envelope , in oxidative stress, and in metal homeostasis and resistance.

Resistance mechanisms against metals are not well characterized for LAB reviewed in reference 26 , and most available information concerns pathogenic streptococci. This allows transcription of the cop operon The Lc. Recently, a different copper resistance mechanism was described for S.

This organism lacks a CopZ homolog. Instead, the resistance system consists of the membrane-bound Cu chaperone CupA, a copper exporter CopA , and a CopY-type regulator The copY , cupA , and copA genes are arranged in an operon whose expression was shown to be induced by Cu and repressed by CopY Bacteria require zinc, but an excess of zinc has toxic effects.

Regulation in Lc. Purified ZitR is a dimer with up to two zinc ligands per monomer. TetR proteins constitute a widespread family of OCSs regulating many aspects of bacterial physiology and interacting with a vast array of ligands Resistance against zinc in S.

Despite extensive studies on other TetR regulators, metal-binding TetR regulators are quite uncommon in bacteria and await further characterization Cytoplasmic manganese plays a significant role in the protection against oxidative stress in LAB Biochemical studies of the S. Later studies showed that SloR controls a large regulon, acting both as a repressor and as an activator 69 , The homologous regulator MtsR of S. MtsR also represses the dnaK operon, suggesting that it mediates the response of dnaK to heat shock Although the oxidative stress response has been researched extensively in LAB, the regulatory mechanisms involved are still largely unknown Although most members of this family are involved in metal homeostasis, PerR is a sensor of H 2 O 2 This has been documented for E.

Evidence suggests that PerR regulons may vary in different streptococcal species. Bacterial genomes contain eukaryotic-type kinases that are mainly responsible for serine and threonine phosphorylation. A number of these have been characterized for some streptococci and E. The S. Indeed, S. A recent study revealed that StkP requires the GspB protein for proper localization in the cell septum and for autophosphorylation and subsequent phosphorylation of its substrates RitR plays a key role in the response to iron and oxidative stresses by regulating the expression of the peroxide resistance protein Dpr and of iron uptake systems The E.

Bacteria possess two main routes to sense a sudden temperature change and to transmit the information. First is the evolutionarily conserved response to the heat-induced accumulation of denatured proteins, and the second is the direct sensing of temperature changes through primary thermosensory structures, such as DNA, RNA, proteins, or lipids, which either have a direct effect or lead to the activation of signal transduction pathways It has been demonstrated for several CtsR proteins, including that of Lc.

The temperature sensitivity of CtsR is adapted to the specific living conditions of different low-GC Gram-positive bacteria The thermosensor region is a highly conserved tetraglycine loop within the winged helix-turn-helix domain.

This conserved region, which possesses high conformational entropy and therefore displays decreased thermostability, senses specific temperature shifts and regulates gene activity, whereas the more flexible regions of CtsR are responsible for adaptation to host-specific temperatures Accumulation of the alarmone p ppGpp in the cell triggers the stringent response SR , a highly conserved bacterial stress response originally defined as a response to amino acid starvation but nowadays recognized as being triggered by a wide range of environmental stress conditions Fig.

The SR induces large-scale transcriptional alterations that ultimately lead to a physiological shift to a nongrowth state. An evolutionary analysis of the RSH superfamily showed that members of the Lactobacillales encode anywhere from one O. The SR has been implicated in acid stress resistance in Lc. A number of studies have shown that RSH is mainly involved in the classical SR, whereas the small synthetases may play different roles.

There is also evidence indicating that RelP and RelQ may have distinct and specialized functions in S. It remains to be established in full detail how p ppGpp modulates cell physiology in LAB. Interestingly, growth inhibition by GTP stress also occurs in E. The mechanisms of the regulation of expression of p ppGpp-producing enzymes in LAB are largely unknown. This indicates that RelRS is involved in the regulation of p ppGpp metabolism The environmental signals to which RelRS responds have not been determined, although it has been suggested that RelRS may sense oxidative stressors or by-products of oxidative metabolism Expression of relPRS is regulated by the MarR family transcriptional regulator RcrR, which is involved in stress tolerance and competence, although the signals to which RcrR responds remain unidentified , There is some evidence suggesting that nucleotide pools may be involved in the modulation of p ppGpp production.

Inactivation of the purine nucleoside phosphorylase DeoD in S. Triggering of the response is regulated by the concerted actions of the repressor LexA and the activator RecA, resulting in the induction of expression of proteins involved in DNA repair A homologous system was subsequently characterized in Streptococcus uberis , in which a gene cluster consisting of hdiR , umuC , and two uncharacterized genes was identified A recent study showed that a LexA-like transcriptional regulator of S.

In contrast to the typical SOS response, activation of S. In streptococci, such as S. It now appears that it is caused by the chromosomal location of early competence genes. Slager et al. This results in a higher copy number of genes close to the replication origin, including the comCDE operon. The increase in gene dosage is sufficient to trigger the competent state The origin-proximal location of early com genes thus constitutes a sensory mechanism that enables cells to activate competence in response to antibiotics interfering with DNA replication Intriguingly, competence in this organism is regulated by the comRS genes, which are located far from the origin of replication Very little is known about their role in LAB physiology.

In recent years, c-di-AMP has been identified as a major second messenger reviewed in reference Inactivation of a gene encoding a DAC homolog in S. On the other hand, inactivation of the Lc. Bacterial quorum sensing regulates a number of cellular processes, including biofilm development, conjugation, competence, bacteriocin production, and pathogenesis. Furthermore, evidence suggests that it also plays a role in stress responses in oral streptococci, where production of competence-stimulating peptides is stimulated by stress conditions Acidic conditions or exposure to spectinomycin increased the expression of the competence-stimulating peptide-encoding gene comC in S.

Two kinds of signaling molecules have been identified in quorum-sensing systems in LAB: the autoinducer AI-2 and small pheromone peptides. AI-2 is a furanosyl borate diester produced and recognized by a wide variety of bacteria for a review, see reference AI-2 is produced from S -ribosylhomocysteine by the S -ribosylhomocysteine lyase LuxS, which is present in most Lactobacillales species, and a number of studies have shown that many LAB respond to AI-2 see the references in references and Inactivation of luxS in S.

It still remains to be established clearly, however, whether AI-2 plays a relevant role in stress responses. The role of small pheromone peptides or autoinducing peptides AIs in intercellular communication of LAB is better characterized. AIs can be detected at the cell surface by specific TCSs or recognized by intracellular receptors after internalization. An example of a pheromone peptide is the previously discussed CSP regulating competence in S.

The second type of pheromone peptides is exemplified by the ComRS system of streptococci from the bovis, pyogenes, and salivarius groups , The involvement of competence quorum-sensing systems in stress responses is discussed above. LAB are subjected to marked metabolic perturbations under environmental stress conditions Fig.

As a consequence of stress, cells lower their metabolic activities, which decreases energy production and the generation of a proton motive force PMF and alters growth and viability 17 , Metabolic adaptation is crucial for survival because it stimulates the production of additional energy and lowers the stress level, e.

The extent of metabolic perturbation and the main routes to reprogram pathways responsible for substrate catabolism in order to adapt to environmental stresses vary between LAB species Schematic representation of changes of the carbohydrate metabolism, glycolysis, and fate of pyruvate in lactic acid bacteria. Under environmental stress conditions, LAB change metabolic and energy fluxes, modify the rate of growth, and adapt the metabolism of carbon sources to the new environment by modifying the synthesis of enzymes and metabolites Environmental stresses inhibit the glycolytic pathway of Lc.

Similar metabolic perturbations are common in other LAB. Consequently, the ability of LAB to efficiently transport and metabolize carbohydrates and other carbon sources, such as malate and citrate, under environmental stress conditions is crucial for growth and persistence. LAB express numerous proteins responsible for carbohydrate transport and utilization. Proton-coupled active transport by proteins from the major facilitator superfamily MFS , the glycoside-pentoside-hexuronide GPH superfamily, and the ATP-binding cassette ABC superfamily and by group translocators, such as the phosphotransferase system—glucose-fructose-lactose PTS-GFL superfamily, is used most frequently LAB modulate the synthesis of specific transporters depending on the type of carbohydrate available , Under acid stress conditions, Lb.

Under the same conditions, an acid-resistant mutant of Lb. Under optimal conditions and when cells are growing in glucose-rich media, HPr inhibits PEP-PTSs for carbohydrates other than glucose, preventing their transport into the cell. In addition, glycolytic intermediates e. The resulting P-Ser-HPr interacts with the global transcriptional regulator CcpA, and the complex prevents the catabolism of carbon sources other than glucose, the preferred sugar in most bacteria, by binding to the catabolic repression element cre upstream of the responsible genes and shutting them down.

Under low-pH conditions, some LAB increase pyruvate kinase activity, which may accelerate the depletion of fructose-1,6-bisphosphate, relieving CcpA repression and allowing the use of alternative carbon sources Acid stress causes intracellular acidification, which decreases the activity of cytoplasmic enzymes Transcriptomic and proteomic studies have highlighted that many LAB enhance the levels of glycolytic enzymes under acid, thermal, and osmotic stresses, but without increasing the synthesis of lactic acid , The level of lactate dehydrogenase Ldh , which is responsible for the synthesis of lactic acid from pyruvate, markedly decreases.

Pyruvate oxidase and phosphate acetyltransferase, used to synthesize acetyl-coenzyme A acetyl-CoA , are induced in Lactobacillus delbrueckii subsp. Acetyl-CoA is rerouted toward the biosynthesis of fatty acids instead of butanoate , , which may enhance the rigidity and impermeability of the cytoplasmic membrane , Changes in pyruvate metabolism were also observed in S.

Despite having a fermentative metabolism, several LAB species possess genes that code for a respiratory electron transport chain Compared to when they are fermenting, LAB display an increase of extracellular pH and biomass under energetically favorable respiration conditions, that is, in the presence of oxygen, heme, and menaquinone , — Fig.

Two molecules of NADH, produced from the oxidation of glyceraldehydephosphate, are reoxidized through the action of Ldh. Thus, the production of lactic acid is reduced and the glycolytic flux is redirected toward production of acetate, ethanol, acetoin, diacetyl, and CO 2. The consumption of oxygen reduces its cytoplasmic concentration and limits the oxidative stress.

Hydrogen peroxide is produced under these circumstances, which may affect the growth rate of Lc. The regulation of respiration in Lc. In glucose-rich media, CcpA represses heme intake and cells grow through fermentation, synthesizing only lactic acid Fig. As glucose is consumed, heme intake occurs and Lc. The shift toward the respiratory pathway produces metabolic changes, which mainly involve pyruvate oxidase POX activity for converting pyruvate into acetyl-P.

The respiratory metabolism of Lc. LAB decrease the synthesis of lactic acid and ethanol under oxidative stress. Diacetyl reductase DAR converts acetoin into diacetyl Depending on the cellular redox state, DAR may also catalyze the reduction of acetoin into butanediol. Under carbohydrate starvation conditions, LAB such as Lc. They shift the physiological state by lowering the synthesis of DNA and proteins and achieving the stationary phase of growth.

At the beginning of starvation, cells accumulate glycolytic intermediates PEP, 3-phosphoglycerate, and 2-phosphoglycerate. Under lactose starvation conditions, Lb. An increase of the catabolism of carbon sources e. This mechanism allows cells to survive without carbohydrates Under long-term starvation conditions, repression of PEP-PTS transporters for carbohydrates and of glycolytic enzymes is mainly due to derepression of carbon catabolite repression CCR mediated by CcpA.

Species belonging to the Oenococcus , Pediococcus , Lactococcus , Lactobacillus , and Leuconostoc genera and oral Streptococcus species use the MLF pathway to enhance survival under environmental stress conditions, such as low pH, a high concentration of carbohydrates, or starvation. This pathway involves the decarboxylation of l -malic acid to l -lactic acid and CO 2. The CO 2 produced by MLF is partially used by carbonic anhydrase to form bicarbonate, further increasing the cytosolic pH.

MLF has also a protective role for S. Under acid stress conditions, lactococci and thermophilic lactobacilli metabolize citrate, producing pyruvate. Citrate metabolism requires the following three steps: i citrate intake via permease CitP , ii formation of oxaloacetate from pyruvate through the activity of citrate lyase CL , and iii decarboxylation of oxaloacetate into pyruvate via oxaloacetate decarboxylase OAD.

The end products are acetic acid, CO 2 , and pyruvate. As described above, the environmental conditions determine the fate of pyruvate. Because of the increased pH in the cytoplasm and extracellularly, LAB take advantage of the cofermentation of citrate with other carbohydrates, such as lactose or glucose. The main protective effects of cofermentation differ among LAB.

In the presence of glucose and citrate, homofermentative and facultatively heterofermentative species e. Each mole of citrate produces pyruvate and, subsequently, acetoin, butanediol, and diacetyl, which protect the cell against acid stress. Furthermore, Lc. One additional mole each of acetic acid and ATP is synthetized for each mole of citrate metabolized. The metabolism of nitrogen, which includes proteolytic and urease systems, plays a pivotal role in LAB growth and survival.

The regulation of nitrogen metabolism occurs mainly via GlnR and CodY , Under conditions of high nitrogen availability, GlnR controls the import of nitrogen-containing compounds and the synthesis of intracellular NH 3. On the other hand, CodY interacts with its BCAA coeffector to control amino acid biosynthesis, transport systems, and peptidases. Genomes of LAB encode extracellular or cell wall-associated proteinases that hydrolyze environmental proteins into oligopeptides, as well as specific transport systems to take up oligo-, di-, and tripeptides and FAA from the environment.

Intracellular oligopeptidases and peptidases hydrolyze oligopeptides and peptides into smaller peptides and FAA. LAB modify proteolysis as a response to environmental stresses. Under acidic conditions, they reduce the abundance of amino acid transporters, and FAA become limiting.

Consequently, many lactobacilli increase the activities of peptidases, such as PepN and PepO , , , The proteolytic system of LAB is also influenced by carbon metabolism VBNC cells of Lc. Much more needs to be uncovered regarding the mechanisms of carbon and nitrogen coregulation. CcpA forms complexes with CodY in B. Recently, it was shown that the aminotransferase IlvE of S.

The metabolism of FAA not only contributes to flavor development in fermented foods but also is crucial for LAB stress resistance. It allows the production of precursors for the biosynthesis of amino acids, fatty acids, nucleotides, and vitamins, generates energy under starvation conditions, and increases the intracellular pH Fig. Deamination and decarboxylation of FAA are the main routes that enhance LAB growth and survival under environmental stress conditions.

Amino acid decarboxylation leads to the conversion of an amino acid into the corresponding amine and CO 2. This reaction is coupled to the exchange between substrate and product e. The translocation is driven by the opposite gradients of substrate and product over the membrane, which are maintained by decarboxylation.

These combined activities generate a PMF because proton consumption during decarboxylation leads to an increase of the intracellular pH Several LAB genomes e. The main FAA pathways induced under environmental stress responses are discussed in the following paragraphs.

All genetic components of this pathway are present in LAB genomes Nutrient sources e. The ADI pathway of Streptococcus rattus is subject to substrate induction and catabolite repression The NH 3 produced alkalizes the cytoplasm, thus increasing acid resistance , In addition, the extra ATP produced via the ADI pathway enhances cell survival after carbohydrate depletion , while the intermediate carbamoyl phosphate is used to synthesize pyrimidines. The intergenic regions of the arcABC genes have several cis -regulatory element sites, which is suggestive of direct regulation of the ADI pathway through CcpA , The arcABC genes of Lc.

This pathway allows the hydrolysis of agmatine into putrescine, NH 3 , CO 2 , and ATP and is a response mechanism to acid and starvation stresses. It represents a potential health risk for humans due to the synthesis of putrescine, a biogenic amine. The AgDI pathway of S. GAD catalyzes the decarboxylation of glutamate into GABA, a bioactive compound that acts as a neurotransmitter in humans.

The uptake of glutamate and the expulsion of GABA are mediated by the electrogenic antiporter, which, together with the decarboxylation of glutamate, produces PMF. Three decarboxylation-antiporter cycles are needed to synthesize 1 mol of ATP. Concomitantly, the extracellular pH increases slightly due to the exchange between glutamate and the more alkaline compound GABA. The uptake of aspartic acid and the expulsion of alanine are mediated by an electrogenic antiporter, which, together with the decarboxylation reaction, produces PMF This pathway has been described for Lactobacillus sp.

M3 , Lactobacillus buchneri , and Lb. Other LAB, including Lb. The exact mechanism of regulation of the AspD pathway is presently unknown. In these organisms, hdcA forms an operon with a downstream gene hdcB of unknown function. HDC activity is regulated via the cytosolic pH. The enzyme is active at low pH, but a neutral or alkaline pH disrupts its substrate-binding site. The survival of Lc.

BcaT and aromatic amino acid aminotransferases AraT play a pivotal role in the regulation of the proteolytic system of Lc. BcaT is repressed by glucose, which confirms that a link exists between carbon and nitrogen regulation. The hydrolysis of urea enhances survival of LAB under acid stress conditions. Among LAB, the urease system has been studied mainly in S.

The urease system produces NH 3 and CO 2 from urea, thus protecting cells against acid stress. CO 2 serves in several anaplerotic reactions that are responsible for the biosynthesis of amino acids and nucleic acids. The urease operon of the oral LAB S. The urease system is induced when S. The expression of the urease operon in S.

I is mediated via CodY Inorganic polyphosphate poly-P is a polymer of phosphoanhydride-linked phosphate residues that can be found as chains of a few to thousands of residues in living organisms In bacteria, the accumulation of poly-P is dependent on the activity of polyphosphate kinase Ppk; EC 2.

The hydrolysis of poly-P is catalyzed by exopolyphosphatases Ppx; EC 3. Lactobacillus organisms, such as Lb. The ability of lactobacilli to accumulate poly-P is correlated with the presence of ppk genes in the strains and the availability of phosphate in the medium , However, Lb. Poly-P has a protective role against oxidative stress in bacteria, where it acts as a chaperone preventing the aggregation of damaged proteins The synthesis of poly-P in Escherichia coli is regulated by hypochlorous acid Poly-P is also involved in the regulation of general stress resistance pathways in different bacteria , but more research is required to fully understand its role in LAB Some stresses are imposed on cells through specific molecules, such as protons, ROS, metal ions, and salts such as NaCl.

LAB have developed various mechanisms to directly counteract the negative effects of these molecules Fig. Consequently, toxic molecules are not always associated with a single type of stress. Schematic representation of the main molecular mechanisms preventing macromolecules from being damaged in LAB. SOD, superoxide dismutase. Among the many mechanisms that LAB species employ to counteract stress damage, the F-ATPase system deserves attention as a system that protects against protons.

The F-ATPase acts in parallel to the above-described metabolic adaptations that result in alkalization of the intracellular and extracellular environments. F-ATPase is a reversible enzyme that can also synthesize ATP by using protons that flow from the environment into the cell. This reaction requires ATP because the expulsion of protons from the cytoplasm to the outside environment takes place against an increasing proton gradient that is a consequence of the synthesis of lactic acid during carbohydrate fermentation Distinctly aciduric oral streptococci, such as S.

The capacity to pump protons out of the cell under acidic conditions protects important glycolytic enzymes and allows long-term survival Lactobacilli increase the synthesis of one or more of their ATP synthase components e. The F-ATPase complex not only is involved in the maintenance of cytoplasmic pH homeostasis but also responds to bile salt exposure , — It was noticed that in Lb.

Oxygen per se does not damage cells, but during the operation of metabolic pathways it is partially reduced to water, leading to the synthesis of ROS, including superoxide anion radicals, hydroxyl radicals, and H 2 O 2. These display a considerable oxidizing potential and are highly toxic to cells , As detailed above, Lc.

Superoxide anion as well as hydroxyl radicals are produced during the Fenton reaction , Metal ions can catalyze certain reactions leading to the formation of ROS, which is in fact considered one of the underlying mechanisms of metal-induced stress. Iron, copper, and other redox-active metal ions exert their effects by stimulating the Fenton reaction. At the molecular level, ROS can react with macromolecules, such as proteins, lipids, and nucleic acids.

Superoxide anions have a limited oxidizing potential and may attack compounds such as polyphenols, ascorbate, and catecholamines The damage caused by the hydroxyl radical, which is the most destructive oxidative agent, includes strand breakage and chemical modification of DNA , Bacteria such as Lc. Notably, induction of genes encoding ROS-protective enzymes is growth phase dependent and confers multistress resistance , NADH peroxidase activity is rather low in Lc. It was recently proposed that metabolically synthetized pyruvate reacts nonenzymatically with residual H 2 O 2 to produce water They catalyze either the two-electron reduction of oxygen to H 2 O 2 performed by Nox1 or the four-electron reduction of oxygen to water AhpCF and thereby provide a very efficient enzymatic defense against oxidative stress Another key player in resistance to oxidative stress is SOD, which eliminates superoxide anion radicals.

Bacteria possess different types of SOD enzymes depending on the bivalent metal cofactor with which the enzyme is coupled, i. Analysis of an Lc. An alternative strategy to counteract superoxide anion radicals is by the production of high levels of intracellular glutathione When this pathway is impaired by restricting access to these nutrients, extracellular superoxide is generated by partially reduced demethylmenaquinones upon exposure to oxygen Thus, E.

Bacteria can also cope with oxidative stress through catalase, an enzyme catalyzing the detoxification of H 2 O 2. The following three classes of bacterial catalases are currently recognized: monofunctional catalases, catalase-peroxidases, and manganese catalases pseudocatalases The monofunctional catalases and the catalase-peroxidases contain heme as a prosthetic group.

The monofunctional catalase specified by the genome of E. In order to retain the low intracellular redox potential required to keep proteins in their reduced form, the presence of glutathione as well as the thioredoxin systems is crucial Members of the genus Streptococcus contain glutathione. They either synthesize glutathione or import it from the medium In addition, streptococci encode a glutathione reductase, which catalyzes the reduction of glutathione disulfide that is produced upon oxidation of glutathione by NADPH The glutathione reductase gene in S.

This relatively large genetic repertoire emphasizes the importance of such enzymes in oxidative stress resistance for a review, see reference Adaptation is also part of the survival strategy of LAB to cope with oxidative stress. Upon exposure to sublethal concentrations of H 2 O 2 , Lc.

PoxB utilizes oxygen to convert pyruvate to acetyl-phosphate Fig. Detoxification of the latter molecule is subsequently done by NADH peroxidase Notably, PoxB activity is induced by oxygen or H 2 O 2 and repressed by glucose In silico analyses of Lb. This is suggestive of regulation of poxB transcription in response to oxidative stress. Other specific mechanisms involved in the adaptation of S. Genome analysis of S. The mechanisms that evolved in LAB to overcome oxidative stress seem to overlap, and thus cells subjected to a specific stress condition can trigger different stress responses for a review, see reference The cAMP receptor protein, the fumarate and nitrate reduction regulator FNR , and FNR-like proteins Flp are part of a superfamily of proteins that encompasses structurally related transcription factors implicated in the control of stress responses As mentioned above, such proteins display structural features allowing transmission of environmental and metabolic signals to the cell and which may trigger general and specific responses to various stressful physiological conditions In Lb.

They are located in two paralogous operons in which the proximal genes are predicted to encode metal ion transport systems It was postulated that Lc. Many regulators in streptococci modulate the interplay between oxidative stress and iron homeostasis, virulence, and competence Iron is considered a key element in regulatory proteins sensing redox modifications in the presence of oxygen This is illustrated by an S. In fact, control of iron homeostasis is considered to be coupled to ROS protection in streptococci Also, the S.

Its involvement in the regulation of proteins related to oxidative stress was revealed when the rgg gene was inactivated Inactivation of an Rgg-like protein of S. Osmotically active compounds serve a dual function in osmoregulation. In addition to maintaining a stable cell turgor, they also exert protective effects on biomolecules in vitro during stressful challenges. Various osmotically active compounds, such as proline and betaine, protect soluble and peripherally membrane-bound proteins from the denaturing effects of modifications in intracellular ionic strength and water availability.

Osmotically active compounds can interact with proteins in several different ways, such as preferential exclusion, water replacement, hydration force, and vitrification of sugars Betaine transport activity in LAB, such as Lc.

Other osmotically active compounds, such as the disaccharide sucrose, protect Lc. The protective effect of ionic solutes has been ascribed to the intracellular accumulation of compatible solutes as a response to osmotic stress. Thus, ionic solutes e. A large number of bacterial ABC-type drug export systems have been characterized Most of these microbial drug extrusion systems are specific chemical or multidrug resistance MDR systems that remove various substances from the cytoplasm or membrane of a cell Notably, LmrA expressed in human or insect cells efficiently exports drugs out of these cells and complements a lack of expression of the human MDR1 P-glycoprotein A homolog of lmrA with highly significant structural and functional similarities to its lactococcal equivalent has been identified in O.

In bacteria, as in all living cells, the cellular proteome is in a constant state of flux. Protein quality control, including refolding or degradation of damaged proteins, plays an indispensable role under both stressed and nonstressed conditions.

Similarly, the catalog of cellular proteins is continuously being modified to meet the challenges arising from changes in the environment. The synthesis of chaperones and proteases is quickly induced under stress conditions to combat the potentially deleterious aggregation of denatured proteins Fig.

While the task of chaperones is to protect functional proteins and to refold misfolded ones, proteases provide the last line of defense by removing irreversibly damaged proteins, after which the amino acids are recycled.

Under changing conditions, proteolysis also removes functional proteins for regulatory purposes or simply because they are no longer needed. It is obvious that the balance between refolding and proteolysis, including the highly specific recognition of substrates, is crucial to avoid wasteful destruction , Schematic representation of the main molecular mechanisms repairing damaged macromolecules in LAB. Chaperone proteins recognize stretches of hydrophobic amino acids typically exposed by nonnative proteins and bind to these regions to prevent aggregation.

They can also actively assist protein folding in an ATP-dependent manner. One well-conserved bacterial chaperone system that can efficiently refold misfolded proteins is that of DnaK Hsp A deletion in dnaK resulted in a thermosensitive phenotype for Lc.

DnaK is essential in S. The inability of DnaK proteins of S. Since only a few reports deal with DnaK structure or biochemical function in LAB, several fundamental questions remain to be clarified. For example, to date, the cochaperone-dependent folding activity of DnaK has not been demonstrated for any LAB.

GroES is the most abundant protein in B. The Clp machinery is probably the main system for general protein turnover in LAB, as is the case in other low-GC Gram-positive bacteria The central proteolytic core consists of 14 ClpP subunits stacked back to back in two heptameric rings, forming an internal chamber that encapsulates the active sites of the ClpP peptidases The peptidase multimer associates with one or two hexameric rings of Clp ATPases to gain proteolytic activity.

The Clp ATPases, together with the proteolytic core, are responsible for substrate recognition, unfolding, and translocation into the ClpP degradation chamber. In the absence of ClpP, threading of substrates through the Clp ATPase ring structures leads to disaggregation and refolding of protein substrates, which are activities typical of molecular chaperones ClpE is absent in E.

With the exception of clpX , the genes encoding the Clp family of proteins are not essential for most bacteria, including LAB , which has facilitated the construction of knockout mutants and the study of the physiological roles of Clp proteins in a variety of LAB species. In contrast, ClpX is not essential in S. Phenotypic studies with mutant strains lacking functional ClpP, ClpL, ClpE, or ClpB revealed the central role of these proteins under stress conditions and, for pathogenic LAB species, in virulence , , — In contrast to the situation in B.

Phenotypic characterization of an S. The alternative sigma factor ComX, which drives the expression of competence genes, was shown in S. This task is performed by ClpEP in S. The primary stress induction of molecular chaperones and Clp proteins in LAB occurs at the level of transcription and is regulated mainly by the transcriptional repressors CtsR and HrcA. The B. One of the repressors is absent in some LAB species, and a single regulator controls expression of both regulons The expression of S.

The HrcA and CtsR regulons may partially overlap in several other members of the Streptococcaceae , Sequence analysis of Lb. An example of reductive evolution of stress repressors with a different outcome is provided by Lb. The HrcA regulon in these species has expanded to include clp genes, in addition to genes encoding the groEL and dnaK operons.

According to studies of other organisms, HrcA is prone to aggregation, a characteristic feature that is also central to its autoregulation HrcA is thought to be released from ribosomes in an inactive form. Thus, protein stress directly affects HrcA activity.

Molecular studies have revealed several differences in the mechanisms of regulation of CtsR function between LAB and other bacteria. McsB is not involved in the regulation of CtsR activity during heat stress in vivo , and CtsR itself acts as an intrinsic thermosensor whose DNA-binding activity gradually decreases with increasing temperature.

The CtsR regulon is also derepressed under several environmental stress conditions other than heat shock, such as acid stress and oxidative damage. This implies that alternative mechanisms for CtsR inactivation must exist. A recent report provided evidence that S. The FtsH proteins of Lc. Inactivation of ftsH resulted in salt-, heat-, and cold-sensitive phenotypes of Lc. The mechanisms controlling ftsH expression in other LAB have not been studied. HtrA high temperature requirement protein A , also known as DegP, is involved in degradation of extracellular polypeptides and misfolded proteins Fig.

HtrA contains a chaperone and a proteolytic domain. An Lc. The htrA gene has been identified as a heat shock gene in Lactobacillus helveticus , and elevated HtrA levels have been detected in Lb. However, the regulatory mechanism behind the stress-inducible expression of htrA in LAB remains to be elucidated. The members of the sHSP family function as folding chaperones, assisting the protein folding process by stabilizing unfolded or partially folded proteins without actively promoting their remodeling Furthermore, sHSPs appear to play an important role in maintaining membrane integrity, especially under stress conditions Lo18 has been suggested to play a dual role in O.

A sudden temperature downshift induces a cold shock response, during which growth of cells is first arrested for a period before reinitiation at a new and much reduced rate. The cellular effects of cold shock include the following: i a decrease in membrane fluidity, ii a reduced efficiency of mRNA translation and transcription through stabilization of secondary structures of DNA and RNA, iii ineffective or slow protein folding, and iv hindered ribosome function In cold-shocked E.

Members of the CSP family enhance the expression of cold shock-inducible genes by destabilizing nonproductive secondary structures in mRNA at low temperatures and therefore are often called RNA chaperones While cold and heat shocks are considered highly separate processes in E. Cold shock of S. Furthermore, the clpL knockout mutant of S. Transcriptional analyses have indicated that clpX is induced by both heat and cold in Lc. The proposed cross talk between cold and heat shock responses in LAB is further supported by the improved cryotolerance seen after heat treatment of Lb.

Similar findings may link acid and cold stress responses in LAB; for Lb. Spx is a global transcriptional regulator of oxidative stress in several Gram-positive bacteria and plays important roles as both a positive and a negative modulator of gene expression Studies of B. DNA microarray work with spx mutants confirmed the function of Spx as a major oxidative stress regulator , Streptococci harbor two bona fide Spx regulators, whereas enterococci have only one , — According to the current DNA damage paradigm, cells undergo an inducible process called the SOS response during genotoxic stress.

This response triggers the expression of genes involved in DNA replication, repair, and mutagenesis The SOS response is repressed under physiological conditions by binding of the LexA repressor protein to operator sites in the promoter regions of SOS genes, including its own gene, lexA While the bacterial DNA damage response is one of the most extensively studied stress responses, it has not been studied widely in LAB. The uvrA gene, which encodes a nucleotide excision repair protein and is a typical member of LexA regulons, has been characterized for S.

Other DNA repair activities that have been implicated in the resistance of S. LAB have cell envelopes that are typical of Gram-positive bacteria. The cell wall itself comprises a PG sacculus that is usually decorated with lipo teichoic acids, polysaccharides, and proteins for a review, see reference The cell wall provides a point of non covalent attachment of important extracellular enzymes and external structures, such as pili and flagella It also participates in processes of adhesion, cell aggregation, and cell division Most importantly for this review, the cell envelope acts as a diffusion barrier between the cell and its surroundings.

It is crucial for maintaining the integrity and shape of the cell and for resisting changes in osmotic pressure. Many of the environmental or technological stressors that LAB can encounter have an impact on the cell envelope. This is why it is very difficult to arrive at a strict definition of cell envelope stress, as regulator systems that respond to stresses exerted on or sensed at the cell envelope often also play a role in the normal physiology of the cell The cell wall is a major target in our fight against pathogenic bacteria, including pathogenic LAB.

Cell wall synthesis can be inhibited by antibiotics that block certain enzymatic steps or capture one of the precursors Antibiotics such as vancomycin, lantibiotics, or bacitracin target the translocation via lipid II of PG precursors across the cytoplasmic membrane lipid II cycle Vancomycin binds to the d -alanyl— d -alanine ends of PG precursors, thereby inhibiting the formation of mature PG Inducible vancomycin resistance was first described for clinical isolates of enterococci , So far, eight enterococcal van gene clusters vanA to - E , vanG , vanL , and vanM have been described, and they encode ligases required for the synthesis of either d -Ala— d -lactate or d -Ala— d -Ser PG precursors for which glycopeptides, such as vancomycin, have low affinity , The molecular mechanisms of regulation of vancomycin resistance gene expression have been reviewed thoroughly and are not discussed here.

The specific ligand recognized by the VanS sensor proteins has not yet been determined unequivocally. Two distinct models have been proposed: direct induction, in which the sensor kinase binds the antibiotic, and indirect induction, in which the sensor kinase binds a cell wall component that either is an intermediate of cell wall biosynthesis or accumulates as a result of antibiotic action Lantibiotics, such as nisin produced by Lc.

Nisin binds lipid II and thus has a double effect on the cell: it inhibits PG synthesis, and it forms pores in the cytoplasmic membrane through which essential components can leak out, killing the cell The main resistance mechanism against positively charged molecules, such as lantibiotics, is a change of the overall negative charge of the cell wall by coupling the positively charged amino acid d -alanine to teichoic acids in the cell wall Bacitracin, a cyclic dodecylpeptide antibiotic that blocks the lipid II cycle by inhibiting dephosphorylation of isoprenyl pyrophosphate , is counteracted by the action of specific ABC transporters BceAB see below.

Other mechanisms, such as the synthesis of alternative isoprenyl pyrophosphate phosphatases or de novo synthesis of undecaprenyl phosphate, have also been described , BceRS was first described for B. A remarkable characteristic of these systems is that IM-HKs are unable to function as sensors: this function is provided by the ABC transporter, with which they are usually genetically and functionally associated , These studies revealed remarkable differences in the genetic organization and regulons.

Both TCSs constitute functional units with their corresponding ABC transporters, and transcriptional analyses showed that while TCS09 controlled the expression of the cognate ABC09 transporter, TCS12 controlled the expression of the orphan ABC as well as the dlt operon involved in d -alanylation of teichoic acids and the mprF gene involved in l -lysinylation of phospholipids One of the ABC transporters is essential for sensing bacitracin and eliciting a response A later study identified four additional genes that belong to the MbrABRS regulon, including those for the cell surface antigen SpaP and for MurM and MurN, which are alanine-adding enzymes involved in the branched-peptide PG biosynthetic pathway.

Although most OCSs are predicted to be cytosolic proteins, some may actually be membrane bound The BcrR bacitracin sensor of E. An auxiliary protein is not required for bacitracin sensing by BcrR Deletion of the transmembrane domain abolished DNA-binding activity, suggesting that membrane localization is essential for BcrR activity Binding of bacitracin did not alter the DNase I protection pattern or the apparent DNA-binding constant for binding of BcrR in vitro to the bcrABD promoter, but it did lead to induction of transcription of this promoter in an in vitro transcription assay On the basis of these results, BcrR is proposed to always be bound to its target DNA sequence; binding of bacitracin would induce a conformational change facilitating a productive interaction with the RNA polymerase, through an as yet undetermined mechanism One of the main functions of the cell wall is to maintain cell shape and to counteract the high internal osmotic pressure, which can reach up to 2, kPa in Gram-positive bacteria.

The cell envelope plays a key role in the regulation of osmotic stress, as it is highly permeable to water, while the cytoplasmic membrane acts as a barrier for most solutes. The main response of LAB to osmotic stress is the intracellular accumulation of osmoprotectants, of which the most common are glycine betaine, choline, carnitine, and dimethylsulfonioacetate Due to their reduced biosynthetic capabilities, LAB have to import these compounds from their natural environments, such as milk, meat products, or plant materials , The ability of these osmolytes to reduce the inhibitory effect of hyperosmotic situations has been established for a large number of LAB , — Another mechanism for LAB to cope with osmotic stress is through regulation of the intracellular concentrations of certain amino acids.

Intracellular concentrations of proline, glutamate, and sometimes alanine and aspartate increase when Lb. Some amino acids reduced the impact of osmotic stress on growth when they were added exogenously to LAB cultured in chemically defined media. This was reported for the addition of proline to Lc. Di- and tripeptides generated by the activity of cell envelope-associated proteinases can also act as osmoprotectants in LAB, as reported for O.

LAB species have developed efficient mechanisms for recruitment of osmoprotectants from their environment. Accumulation of osmolytes is mediated by transporters that move solutes across the membrane against their electrochemical gradients in processes that require metabolic energy Interaction of the transporters with membrane lipids determines their activity in response to osmotic changes Betaine and proline uptake in Lc. This operon encodes a transport system of the ABC family that uses two ATP molecules per translocated molecule of glycine betaine OpuA detects changes in the intracellular ionic strength, and its activity depends on electrostatic interactions between lipid head groups and OpuA In particular, sensing depends on the C-terminal tail of the CBS domains, which is negatively charged A model was proposed in which the transporter switches from the active to the inactive state depending on the interaction of the CBS domains with lipid head groups in the membrane.

Repulsion between the charged C-terminal tail and the anionic heads of the membrane lipids counteracts the interaction between the CBS domains and the membrane. In this way, the intracellular ionic strength and the concentration of anionic lipids in the cytoplasmic membrane determine the activation of the transporter A later study showed that the interaction of the CBS domains and the membrane lipids depends on a surface-exposed cationic region in the CBS domain, in agreement with the proposed model A genome-wide gene transcription analysis revealed that Lc.

In Lc. The uptake of osmolytes by Lb. This system has a high affinity for glycine betaine and a low affinity for proline and is expressed semiconstitutively, i. When bacteria are faced with a sudden drop of the external osmotic pressure, they use mechanosensitive channels MSC to counteract the effects. The MSC act as emergency release valves, allowing the expulsion of osmolytes and water.

MSC share an organization of a C-terminal cytoplasmic domain and an N-terminal transmembrane domain formed by symmetrical helices that can change between open and closed states Although their mechanism of sensing is still unclear, it is well known that MSC can sense changes in the tension of the lipid bilayer of the cytoplasmic membrane Most microbes appear to possess members of one or both families of bacterial MSC Although Lc. LAB can counteract several stresses by modifying their cytoplasmic membranes.

The regulation of membrane fluidity in bacteria is a rather complex process that often involves universal protein chaperones, such as GroEL and sHSPs , The fluidity of the cytoplasmic membrane is highly influenced by the ratio of unsaturated to saturated fatty acids, and alterations in membrane fatty acids may represent a generalized response utilized by many bacteria to survive environmental stresses affecting the membrane.

The ratios of unsaturated to saturated and cyclic to saturated membrane fatty acids decreased in a strain of Lb. The decrease in membrane fluidity in Lactobacillus could be compensated by other variations, such as an increase of cyclic fatty acids

HOW TO READ HORSE BETTING BOOKS

The far eastern manufacturers, Sunrace Sturmey-Archer and Shimano, are much more vague on what is an acceptable minimum input ratio. No actual hard numbers are listed in their literature. I contacted technical support here in the U. Shimano stated that with the stock input sprocket chain rings down to 33 teeth are ok. The recommended range listed was 33 to 38 teeth.

This allows an input ratio of as low as 1. If geared this way then high gear is very low even on the 8 speed unit, in the neighborhood of a development of 5 meters. Too low to be practical in my opinion. I have not been able to determine what that is for the 5 speed. For the 8 speed it is either the 25 tooth or 27 tooth sprockets, the only two offered for this hub.

With the low gear on the 8 speed being direct drive this makes sense for their 8 speed but still must subject the internals to very high torque loads. The 5 speed is still basically the same as the old pre Sunrace design so far as I know. Pre Sunrace literature requires a minimum input ratio of 2. I would suspect that use of the 30 to 33 tooth chainring would lower the input ratio, and increase the torque loads the mechanism is subjected to, substantially.

Is anyone aware of redesigns or strengthening to this hubs internals which would allow this change in recommended input ratio? Are Shimano and Sunrace ignoring the more performance oriented users of their hubs and using specifications aimed at those who buy cruiser type bikes? Are their recommendations ignoring more performance oriented riders who can put a much greater strain on a gear hub, risking damaging it? October 22, at pm. December 7, at am. Hi everybody. If anybody has one to sell please send me an email to nexus sportfund.

January 15, at am. I run a Shimano Nexus 7 on on of my winter bikes. I live in Ottawa, Canada, where it can get pretty darn cold, and the hub has been quite reliable. I acquired it secondhand, and having not opened one before I took it to my local shop where the owner has actually worked on these. He said the bearings on one side were shot, but it was otherwise fine.

When converting the bike to this hub I installed as little cable housing as possible using a Sturmey cable housing stop and the little guide wheel , and have had no shifting issues, even at C. And I was fortunate to find a thumbshifter, as using a gripshift with thick winter mitts is not fun.

I have it geared for winter trailer towing, so on flat roads with no towing I cruise in 6th or 7th, tough with a load 1st certainly does see use. But I do dislike the way the shift cable connects to the hub. Just make sure you have a crash guard over the clickbox and no worry of damage.

On the Nexus I was thankful that one can remove the roller brake, as that too was another bolt to undo in the cold. I just got an old SA AG Dynothree hub 3-speed and generator combined , and now need to build it into a bike. February 3, at am. Any word whether this hub will be available aftermarket? I am very much interested in a 3-speed IGhub with a disc mount.

The SRAM website lists it as having a disc brake option…. February 28, at pm. I would guess it is level with the flats but does anyone know for sure? April 7, at pm. Conclusion: the germans are the real hub masters; therefore I would like to test an sram 9spd on a MTB. June 20, at pm. Internal hubs make the most sense in urban riding where quick shifting for acceleration; reliability; efficiency and reasonable cost are more important than having very even, small ratios.

Classic Sturmey-Archer hubs were designed for light weight, unlike most newer hubs. The steel hub shell of one SRAM Spectro S7 I have is monstrously, unnecessarily heavy and the aluminum hub shell of another is still unnecessarily heavy! Also, note that the ratios given on this page are slightly inaccurate. They appear to have been derived from rounded percentage numbers. The actual ratios are fractions which may not round exactly to percentages.

The formula for gearing up is. The formula for gearing down is the inverse,. You may calculate the percentages yourself. The formulas are somewhat more complicated for hubs with compound gearing, but all produce exact results as fractions. Hubs with more sets of gears on the way from the sprocket to the hub shell are generally less efficient, as are hubs lubricated with grease rather than oil. August 31, at am. Love this machine.

Fast acceleration — and as your chart shows — nice intervals on shifts. My guess is my top speed is somewhere around 20mph peddling. So far so good — done about miles on the bike — city riding — and one country trip of 10 miles on steep hilly roads. September 22, at am. I think the even gear spacing comments by Marco is too academic.

Hub designers put in large steps for first and last gear on purpose. If you are going up hill and need a low gear, you would need a really low gear. If you are going down hill and want more speed, you would want a big jump in gear ratio. This is especially true for few gear hubs and cargo hubs.

If the bike is loaded, like propane tank delivery bikers, you would want a really low gear. September 12, at pm. Thank you Marco for an absolutely wondeful site. It provided me with all the information I needed to make an informed decision.

September 28, at pm. The hubs are great but I keep snapping spokes. My tips for building your own wheels… 1 the spoke holes on the rim might be handed i. Yes I got that wrong even chance I suppose 2 lace the wheel 1 cross. I laced 3 cross and this puts a big angle on the nipple end of the spoke because of the diameter of the hub flange.

All my spoke breakages were at the end of the thread. On smaller wheels it is even more important to reduce the number of crosses. Or if you use one of the massive nuvinci hubs! October 21, at pm. Thanks for a great site! Ran on a Nexus-7 hub for a couple of winters. Opened it up and was pleased to see that even with running in rain, snow and brine, it still looked pretty good. External gear shift ring hard to reseat cable clamp when replacing rear wheel.

If shift cable creeps out of adjustment, hub does not shift well until re-centered on mark. Shifting VERY positive, no gaps between gears. Shifting easier than Nexus hub. Maintenance practically 0 oil change. Range of gearing is incredible. Hub runs very smoothly and has never caused a problem. Also the external gear box that I use with disc brakes is extremely easy to disconnect and reconnect when removing the rear wheel.

Shifting still not as smooth as a top end derailler system. Shifter never need to be adjusted as indexing is inside the hub. Oh, I also love that gears run in a linear fashion , forget about little chainring-big chainring and bad gear combinations! January 13, at pm. After decades of derailleur use I got a Carrera Subway with a Nexus 8 and at first found it wonderful, especially being able to change gear while stationary. But, after 18 months: disaster, the inner unit somehow separated from the outer shell and the whole thing ground to a halt.

Only a few parts of the innards were broken but the local cycle shop found that spares are not available and that the whole unit would need replacing at total cost of parts and labour not far short of the cost of another bike! I am reverting to using a derailleur, albeit reluctantly. January 28, at am. I dismantled my Sachs Pentasport last week to investigate a faint bearing grumble. This hub has been in constant use since and has covered a documented kms [in reality probably closer to ].

It has never been dismantled or lubricated in that time, still had the factory grease in it [I am ashamed to say]. After a soak in the ultrasonic bath the problem became apparent. The planetary gear teeth crests had worn to a knife edge and were starting to break down, the chips ending up in the RHS large bearing race. The teeth still had good engagement so polished all the bearing tracks and fitted new balls all round, re-lubed it and put it back together. This hub has no O-rings or seals yet it was free from dust and water.

It has no Sprag clutches yet is almost silent. Requires no special tools to dismantle or assemble. It has never jumped or slipped under load. Gear ratios are spot on. Was inexpensive to purchase. This page needs updating now. Shimano just today, 2 February, has announced their new SG-S geared hub. Estimated price of Euros. The original announcement is on a German language web site. Google for it using the model number I have listed.

February 26, at pm. From the custom rims matching the paint job, to the custom colors, all the way to the 2 toned seats, this bicycle was made for people who want to ride as fly as they drive. Not to mention its sleek geometry makes it ride as comfortable as a Cadillac. Even with a friend on the handlebars, it still handles well. March 4, at am. They seem to be a little cheaper and really sold on the whole drum brake thing.

The plan is to go drum brake front and rear on a Surly Cross Check frame with flat bars…. March 16, at pm. I just want to say thanks so much for this article, and all the comments. March 23, at am. Interesting range of comments. I have been using a Shimano 7-sp hub SG-7R40 for 13 years, 40, km. Although it is as smooth and quiet as ever I cannot recommend it because of the unavailability of spare parts.

The bearings are not well sealed and the right-hand bearing cone has failed for the second time. It seems that most of the hubs on the market now come with twist-grip shifters. For those of us who prefer thumb shifters it would be interesting to hear from anyone who has used shifters of brands or types other than the recommended one. April 6, at am. I have an Alfine Hub that I bought nine months ago.

Yes, I have ridden it through some water totally submerged. But, at the end of last season, it started acting funny. Things can to a head early this riding season. The bike shop told me I totally rusted out the hub. I did have over miles k on it, but I was told it only need maintenance every other year.

Turns out that seems to not be true. I guess I need to learn to take this baby apart and grease it up. Anyone have a suggestion on oil to bathe these things in? I saw some for a certain motor oil somewhere on this site. Anyone tried it? April 14, at pm.

April 15, at pm. April 16, at am. July 30, at pm. Who came first? Sturmey and Mr. February 16, at pm. Ok now back to the late comers to the game? Wonder about the internal torque loading, previously talked about, since My AW3 [15t cog] has input equivalent of a May 15, at am.

October 5, at pm. May 16, at am. PS sorry if this is the wrong section but I find this site really confusing to navigate. June 12, at am. Hi excellent review. But I have also read that a hub might look good on paper but suffer from too much internal friction. I have heard that this can be a real problem in some hubs especially as you move further from the slot. June 12, at pm. I think there was also something more recent in Bicycle Quarterly, but only on derailleur systems.

But for the rest of us it is hard to detect, resulting in less than 1 minute per hour difference in average speed. Rohloff claim that the ease of shifting without thinking with fully sequential gears can compensate to some extent for the theoretical shortcomings such as larger steps between gears. There is definitely something in this, as I often notice when riding with people using derailleur gears that they change gears much less often than I do, despite having closer-spaced gears.

They are inhibited either by the effort of determining what combination of front and back sprockets they want or by fear of throwing a chain. There is an analogy with the choice of transmission in cars. To get round a racetrack in the shortest time you would generally choose a manual transmission, but many people have good reasons for choosing automatics, which are often quicker in everyday driving.

Randal, great thoughts. For city riding the hub wins for most riders particularly because you can change when stopped and also change rapidly from top to bottom in gears especially with twist grip changer. I notice in city riding less skilled riders do indeed ride as if they are on a fixie! July 26, at pm. I suspect that the efficiency loss is much greater than what the marketing spin-doctors want us to know. If on a bicycle we are anywhere near such losses, the inefficiency is enormous.

The larger physical size your gears are NOT the ratio — is the same as for example , the less losses, because the tension point is farther from the pivot point bearings , but the downside is added weight. It would be good to see the internals — that would allow us to at least guesstimate although this would not take into account precision of manufacturing.

Click to access hp I am a former racer, team mechanic, and team trainer. Let me offer an alternate viewpoint on gears in general although this is written more with the 3-speed in mind, it applies to any number of gears :. I partially disagree with you, regarding the steps. For example, a rider in good shape who really can spin, may prefer to have a smaller step on the high gear… or a larger step because they may use it only in exceptions, where they have a strong wind in their back or going down a long hill.

Or they may prefer a larger percentage on the smaller gear, for hills, or a smaller one, if they are a strong rider and hate having to spin to go up one. Hence, for sustained high speed, a lower difference between neutral and high might be more desirable which is opposite of some of the hubs being offered — maybe they had fast downhills in mind and easy uphills when they designed the gearing in at least one of these models.

July 29, at pm. I was really out to lunch on that one, as are all the reviewers… The thing is, in a 3 speed hub, there is only one internal gear, that you inverse, to offer one increase or one decrease in the ratio. But in actuality, we need to use the second gear as our starting point. August 27, at am.

Can I get in Germany market? October 11, at pm. It is too early to comment on reliability, of course, but I can make some initial comparisons with the Nexus 7-speed that I used for the previous 13 years. It is also marginally less convenient to use on my bike than the Nexus because I use drop handlebars and the SRAM shifter is only available as a twist grip. I have no way of proving this. The P5 has a fractionally greater range than the old Nexus 7-speed less than the current one, I believe.

The bigger steps of a five-speed gearbox are to my mind an advantage. Unlike many contributors to these forums, I am not fussy about closely-spaced gears. A couple of minor disappointments with the P5: the chrome plating of the shell was a bit patchy, and the weight was 1. A friend of mine recently bought an S7 on special in America for less than I paid for the P5. He has yet to fit it to his bike.

October 12, at am. The question is how many do I get? I have had a good run out of the Pentasport I referred to above [28 Jan 10]. Was able to get replacement planetary gears for it and then did an km tour around Tasmania in May. As you mentioned, although five speed, the ratios are sensibly spaced. October 16, at am.

The SRAM clickbox seems to work quite well, though it looks a bit ungainly, perhaps. This is an important factor in choosing a brand. My car is 27 years old and I have not yet had any trouble getting parts for it. I was able to acquire the whole carrier with planetary gears for the Pentasport. Digital images indicated it was close.

Only problem was the new carrier was for a coaster brake version and the LH end of the carrier had a reduced diameter with a double helix milled into it. My non-brake version was solid with a circlip to retain the pawl ring.

I turned up a collar with a couple of grub screws and mounted it over the helical spigot to keep the pawl ring in place and it has been there ever since. Was also fortunate to pick up a brand new original Pentasport at a swap meet recently as well.

It has a double shift-lever arrangement which I am not familiar with. Prefer the dual cable thumb shifter but hard to come by these days. My last vehicle was 32 years old when I retired it and I am still running my motorbike after 30 years. This was evident with the desperate stimulus packages that flourished during the GFC. Unfortunately this also happens to be in direct conflict with attempts to look after our planet home. November 9, at pm.

Chris I recently had the axle of my Sachs Super 7 break after approximately 14 years of hard work. The site is in Dutch English under development. Enough of Dutch lessons. They seem to supply globally but shipping costs can be expensive. I should add that I have been a satisfied customer and am not connected with this company. December 6, at am. Noisy at first with noticeable internal drag, but has freed up considerably and is quieter now as things settle down and the grease gets distributed through the workings.

A couple of shots of oil through the axle end-hole has quietened it down even further….. The plastic construction may be a blessing in disguise…allowing for the thing to be torn away in the event of a prang rather than damaging the axle. Sort of sacrificial. There was advice in the original Pentasport instructions about leaving it parked in fourth gear to ease the internal springs. Consequently my Alfine gets parked in first gear as does the Spectro.

September 14, at pm. I have two of these hubs, and the second one I purchased had some trouble after about 3 months of daily use. I talked to them their email support is pretty busy, so I called , and they were quite helpful. They were able to send me new internals for my hub, as well as the wrench needed to do the swap. They seemed curious to see what had happened, and it sounded like they may have had a bad batch of hubs.

After about half an hour of work, their wrench and my old hub internals were on the way back to them, and my hub was as good as new. If it happens to you — call them up! December 13, at pm. Or does anyone want to sell a used one they are not useing , I was looking for the N Nuvinci, and anyones insites about this hub. Any and all help will be greatly appreciated. January 26, at am. February 4, at pm. Nice site, interesting review, but why are at least the Sachs three speed hubs Modell 55, , not included?

Something essential is missing here…. March 17, at pm. We found this Stormy Archer 3 speed hub with coaster breaks adult tricycle in a basket. I have already had the hub rebuilt. Can it carry say a person lbs??? What items will I need to get if the sturmey archer option will work. Do I need a grip shifter or can I use a bar end shifter that I have seen advertised below. June 6, at am. Robert, the short answer to your query is that you cannot mix shifters and hubs at will.

The type of shifter available is a significant factor in your choice of hub. What you can consider, though, is a number of hubs with axle widths up to mm or so. July 21, at pm. July 25, at pm. For the 3 speed hubs, it seems hardly worth noting that the Nexus step difference of.

And I have to agree with what others already said about the Sturmey-Archer 8 speed. The hub itself may be junk for all I know, but having a extra big step at each end seems like an advantage to me. At least for recreational purposes. A super low gear to let you fight your way up a hill that maybe for practical purposes you should just get off and push the bike up, and a super high gear for when you want to see how fast you can go down steep hills on the open highway.

You still got 6 evenly-spaced gears for normal riding in the middle. I remember a bike I had with a Nexus 3-speed hub… Top gear was perfect for level ground, but a bit of slope or headwind and I found I could not maintain that gear for long and was forced to go to the middle gear.

Which did make me feel a bit silly for pedalling so fast for the mild conditions. July 29, at am. I agree with you. I own a Terratrike Rover trike, and I find that the big step down to first gear, is almost like having a granny gear. On flat terrain I ride mostly in 6 and 7. I dearly loved the Nexus 3-speed I mentioned before, despite the shift jumps being large. The simplicity of it was really appealing, and for the place I lived then the gear range was perfect.

That would be a huge help for the hilly place I live now… and the 5th gear in there dividing everything up finer would largely solve the step size problem I felt. I think my pea-sized brain could keep track of 5 speeds OK… it really seems like the best compromise between step size, overall gear range, and simplicity to me. Aside from simplicity of operation, one would think having fewer gears would mean each gear set and each pawl etc.

Why no mention of the new Sturmey Archer 8 speed, XRF8 which is a different design than the previous model that had some reliability issues. I have about 5, miles on one on my Trike, and it is the best shifting device I have had, and I have been cycling for over 50 years.

The Sturmey was hard to shift and when shifting from 7 to 8 it felt like it shifted the wrong way, it was not geared low enough for me and was hard to shift down into the lower gears. You had to back peddle to get it to shift. I was just not happy with it. August 13, at pm. September 11, at pm. Which was type of inspiring! Totally unanticipated. September 15, at pm. My low range is about 14 gear inches and the high to high for me to use.

No shifting just adjust as needed to find the sweet spot. I wish I had made the upgrade when I purchased. Would have cost me less and I would have been happier. I agree that Terra Trike needs to take a close look at their components. Next time maybe a Catrike. October 3, at am. Yes, I checked the adjustment and it appeared to be correct. I think the Sturmy is just a crappy hub. I love the Nuvinci and it shifts best while pedaling. Even if it worked correctly the Sturmy was not geared low enough for me.

I need some paper of it the bill and warranty. I would be very thankful. November 5, at pm. Now please, before you in the Spandex crowd Poo-Pa those those of us that simply need to bike commute without having to pedal ALL the time, hear me out and leave the culture of biking out of it please. As we all know gears change everything, and I am one of the few motorized bike builders that does this the right way all the way to the back wheel. For me, it has come down to bicycle drive wheels designed for human power to be able to handle much MUCH more power.

Thanks for the help and if you want to see some examples and even a couple of videos of how much fun these are you can visit my web site. November 8, at am. I know it has been a couple of days but I was hoping someone might have seen this and had an idea for me. November 9, at am. The NuVinci will be the toughest internal hub available.. November 11, at am. Thanks guys. I run a NuVinci on my personal ride and pumping 3. Backing off the power when shifting is nothing, I even do it with the NuVinci.

We are developing a new high torque 5 speed hub but that is slated for production mid If you are interested in this hub please keep in contact. That sounds promising. December 10, at pm. Lower torque at the hub delivering the same power at higher wheel speed might help. December 11, at am. It could work with a rear disc or drum brake however and I see the logic, considering these little I only use human power but I know that Utah Trikes has use the Nuvinci on motorized trikes.

I bet they could give you some hints or solutions. November 26, at am. November 30, at am. Getting ready to order a Hase Kettwiesel Custom. I willuse it for heavy touring and comuting and your info on the rohloff has convinced me to outfit it with one. January 23, at pm. Are there online shops where I can buy hub parts? December 29, at pm. Thank you a bunch for sharing this with all people you actually understand what you are talking about! We will have a link trade contract among us.

January 10, at pm. Everything follows a set of the entirely personal preferences of the author. It would be better just to list the specifications and the gearings, and leave cyclists to make up their own minds, rather than try to impose his prejeudices. January 27, at pm. Spanner48, I agree this review leaves a great deal to be desired, but your question about hub values does not make much of a point, in my opinion.

Those KS and K hubs, etc. And maybe they are rare now because they were less desirable generally when they were originally produced, and sold less. At any rate, the author of this review happens to be advocating close ratios over wide ratios, not the other way around.

January 24, at pm. Can someone perhaps write a review on Joytech 3-speed hubs which are found on new cruiser and city bikes? April 27, at am. Great article! I only offer because I think this is a very useful article that every bike mechanic, professional or amateur, should read! April 30, at am. I would like to start a list of bicycle makers who are using 3-speed hubs.

I find 3 speeds to be the ideal city bicycle. Super sweet looking bike, look it up. May 29, at pm. Thinking more about internally geared hubs, does having the same percentage of increase for each step really make sense? Is that really optimal? That could be useful if you are in a bicycle race and trying to climb a given hill at the best rate possible for your physical condition, but who in such a race is going to be using an IGH?

For real-life use, say climbing a short hill on the way to shopping, you would do just fine staying in the 10 MPH gear, maybe losing 10 seconds on the hill but having a simpler cheaper more durable hub with fewer gears in it. If you are just going through the gears accellerating from a stop on flat ground, the The jumps should be bigger at the low-gear end, and taper down to smaller jumps at the high-gear end.

For the answer, look at derailleur blocks [whether freewheel or freehub]. Some — mostly the s ones — have constant ratio progression. Personally, I prefer the old constant ratio ones. Back in the days, riders would mix and match sprockets to get the gear progression that suited them. November 6, at pm. In my view, if all website owners and bloggers made good content material as you did, the web shall be a lot more helpful than ever before. December 21, at am. November 26, at pm.

I would use a Nexus 8 most likely. February 11, at pm. Can anyone tell me what options are there for mm OLD spacing internal geared hub with 5 or more gears? What brands and model numbers? You could get away with springing the frame out by 2mm, but I would recommend bending the chainstays and checking the axle alignment with a dummy axle to avoid putting any bending load into it when the wheel is installed.

It is also possible to reduce a Shimano Nexus 7-speed from the original mm to mm. I have done this by removing the roller brake from my original hub and using a few parts scavenged from one with a coaster brake. There are different thicknesses of axle bearing locknuts available so you can shave some width from the drive side as well as the left-hand side. The wheel must still be dished, but not by much compared to wheels for most derailleur bikes.

In the case of the SRAM P5 the wheel remains undished if you are using a caliper brake; there is a tiny offset if you use a coaster brake. April 23, at am. Apple and Samsung screens are vibrant and pop out of the bottom. This is not exactly surprising given how much Samsung seems to love thin and cheap-feeling plastic back covers. Your web site provided us with valuable info to work on. You have done a formidable activity and our whole neighborhood can be thankful to you.

May 23, at am. June 9, at am. Roger It is now how do you feel about the Schlumpf and NuVinci? What Schlump did you use and what was the Schlumpf chain ring tooth and NuVinci cog tooth? Think of doing the same. June 10, at pm. I still love the combination. I did change the rear sprocket to a 22 tooth to increase the low end. I may have went too far and may go back off to a The original was an 18 tooth. The Schlumpf is the high speed with a 27 tooth sprocket.

August 6, at am. August 7, at am. August 18, at pm. Can anyone comment on experiences with the Sturmey S2 non-braked 2-speed hub please? I notice the S2C braked version seems to be plagued with problems, but they appear to be associated more with the brake. November 27, at pm. January 9, at pm. February 10, at am. April 26, at am.

My partner and I stumbled over here different website and thought I may as well check things out. Look forward to exploring your web page for a second time. The science behind Adiphene consists of 12 of probably the most power fat preventing ingredients known. Glucomannan has a particular means to soak up water and swell inside the stomach.

In order to energize yourself and improve your outlook you will have to change your diet and get moving. June 17, at pm. Google — Preview — A simple tool, does just as its name says. By using the Internet and search terms, you can find out what is most interesting to your readers and capitalize on those topics by writing about them on your blog and website.

June 21, at am. If you want to know the crystal clear details it is better to view just about any online store web sites explaining the actual product or service information using elements. Adiphene is the newest and fastest weight reduction supplement in market. August 8, at pm. All the best. August 13, at am. My partner and I stumbled over here from a different web page and thought I might as well check things out.

I like what I see so i am just following you. Look forward to going over your web page again. August 17, at pm. This blog was… how do you say it? Finally I have found something which helped me. August 25, at am. Excellent submit. In any event I will be signing up to your rss feed using this program. September 5, at am. Great website you have here but I was curious if you knew of any community forums that cover the same topics discussed here?

If you have any suggestions, please let me know. Bless you! September 20, at am. I hope to offer one thing again and help others like you helped me. September 20, at pm. Download generator now and have a fun. Would you propose starting with a free platform like WordPress or go for a paid option? Any tips? Thank you! My brother recommended I might like this blog. He was totally right. This put up actually made my day. You can not imagine simply how much time I had spent for this info!

What could you recommend in regards to your post that you simply made some days in the past? Any sure? Your help is requested, please. Because, I am re-entering cycling after 40 years being absent; my interest is focused towards recumbent tadpole tricycles. I would like to plan to add later an EcoSpeed mid-mount electric boost motor. Also, I understand that NuVinci N hub is very good, too. Now, throw in the Schlumpf two speed bottom gear and you may understand my angst. So, consider a Steintrike Wild Wave carving recumbent tadpole tricycle to hang all this on.

Will You give me your ideas? One trip is to cycle up and down Mt. Crank arm length? Chain size? Wrist rests? Lights and power sources? I want to hear from you. I use a Nuvinci N combined with a Schlumpf high speed and love it. I replaced the standard 18 tooth gear on the N with a 20 tooth and have plenty of bottom gearing. Utah Trikes does lots of custom configurations and could give you lots of advice on various combinations of hubs, gearing, electric etc.

Hi to every one, the contents present at this website are actually awesome for people knowledge, well, keep up the good work fellows. November 11, at pm. Your opinion on fitting my new recumbent tadpole tricycle with needed equipment is solicited. The Steintrike Wild Wave carving recumbent tadpole tricycle is the trike frame I am considering acquiring. I am pretty sure I will use the mesh seat. I am close to 70 years old, and do not race; but, plan lazily tours.

The Schlumpf two speed bottom gear and Rohloff speed hub coupled with a mid-drive EcoSpeed mid-mount electric boost motor is what I think I want. Please, give me your ideas and opinions. Any suggestions are appreciated. Thanks, Garry. It sounds like a workable combination. You should check Utah Trikes web site. They have done a lot of electric combinations and have experience with the Nuvinci and the Rohloff hubs. They could tell you the pros and cons of each combination. There is lots of information on their web site.

November 14, at pm. Nice info but the commentary is odd at times. Sturmey Archer was certainly not having a bad day. They reached the range of most of the competitors, rollhof notwithsanding, with much finer gear spacing in the cruising gears. This is clearly a gearset made for someone who wants fine cadence control most of the time but still wants the ability to jump up or down to those really low and high gears when needed.

Undeniably believe that which you said. Youur favorit reason seemed to be on the web the simplest thing to be aware of. I say to you, I definitely gett irked while fplks think about issues that they just do not realize about. You controlled to hit the nail upon the top and also defined oout the whole thing withoout having side effect , other folks could take a signal.

Will likely be again too get more. November 22, at am. Normally I do not learn article on blogs, however I would lile to sayy that this write-up very forced me to take a look at and do it! Yourr writing style has been amazed me. February 14, at am. Greetings from Colorado! Anyhow, great blog! I am still reading and re-reading your article to detach for an answer to my question. You have answered so many of my questions well then I am sure my answer will be here.

My question is how high gear do I need? If I go downhill then I want a high gear. June 11, at am. October 18, at pm. I have a Trek Soho that is about 7 years old with the Shimano Nexus 8 speed and a belt drive. Over the summer, a bungee cord got wrapped up between the hub and belt, causing the hub to crack open. I have had it serviced, but the issues with it keep getting worse. The shop has taken it apart and lubed it as well as fixed the drop outs which must have also gotten bent in the bungee episode.

When free-wheeling there is a noticeable vibration through the frame and pedals and the when on a stand, the back wheel slows down quickly after setting it in motion. Also, the shifting is clunky at best. I feel like I have a few options. Replace the guts of the internal hub keeping the old shell2. Build a new wheel around a new hub or 3. Any advice? I do love the Soho — it is such a great workhorse as a commuter bike. March 29, at pm.

April 25, at am. The resulting P-Ser-HPr interacts with the global transcriptional regulator CcpA, and the complex prevents the catabolism of carbon sources other than glucose, the preferred sugar in most bacteria, by binding to the catabolic repression element cre upstream of the responsible genes and shutting them down. Under low-pH conditions, some LAB increase pyruvate kinase activity, which may accelerate the depletion of fructose-1,6-bisphosphate, relieving CcpA repression and allowing the use of alternative carbon sources Acid stress causes intracellular acidification, which decreases the activity of cytoplasmic enzymes Transcriptomic and proteomic studies have highlighted that many LAB enhance the levels of glycolytic enzymes under acid, thermal, and osmotic stresses, but without increasing the synthesis of lactic acid , The level of lactate dehydrogenase Ldh , which is responsible for the synthesis of lactic acid from pyruvate, markedly decreases.

Pyruvate oxidase and phosphate acetyltransferase, used to synthesize acetyl-coenzyme A acetyl-CoA , are induced in Lactobacillus delbrueckii subsp. Acetyl-CoA is rerouted toward the biosynthesis of fatty acids instead of butanoate , , which may enhance the rigidity and impermeability of the cytoplasmic membrane , Changes in pyruvate metabolism were also observed in S.

Despite having a fermentative metabolism, several LAB species possess genes that code for a respiratory electron transport chain Compared to when they are fermenting, LAB display an increase of extracellular pH and biomass under energetically favorable respiration conditions, that is, in the presence of oxygen, heme, and menaquinone , — Fig.

Two molecules of NADH, produced from the oxidation of glyceraldehydephosphate, are reoxidized through the action of Ldh. Thus, the production of lactic acid is reduced and the glycolytic flux is redirected toward production of acetate, ethanol, acetoin, diacetyl, and CO 2.

The consumption of oxygen reduces its cytoplasmic concentration and limits the oxidative stress. Hydrogen peroxide is produced under these circumstances, which may affect the growth rate of Lc. The regulation of respiration in Lc. In glucose-rich media, CcpA represses heme intake and cells grow through fermentation, synthesizing only lactic acid Fig.

As glucose is consumed, heme intake occurs and Lc. The shift toward the respiratory pathway produces metabolic changes, which mainly involve pyruvate oxidase POX activity for converting pyruvate into acetyl-P. The respiratory metabolism of Lc. LAB decrease the synthesis of lactic acid and ethanol under oxidative stress. Diacetyl reductase DAR converts acetoin into diacetyl Depending on the cellular redox state, DAR may also catalyze the reduction of acetoin into butanediol.

Under carbohydrate starvation conditions, LAB such as Lc. They shift the physiological state by lowering the synthesis of DNA and proteins and achieving the stationary phase of growth. At the beginning of starvation, cells accumulate glycolytic intermediates PEP, 3-phosphoglycerate, and 2-phosphoglycerate. Under lactose starvation conditions, Lb. An increase of the catabolism of carbon sources e. This mechanism allows cells to survive without carbohydrates Under long-term starvation conditions, repression of PEP-PTS transporters for carbohydrates and of glycolytic enzymes is mainly due to derepression of carbon catabolite repression CCR mediated by CcpA.

Species belonging to the Oenococcus , Pediococcus , Lactococcus , Lactobacillus , and Leuconostoc genera and oral Streptococcus species use the MLF pathway to enhance survival under environmental stress conditions, such as low pH, a high concentration of carbohydrates, or starvation. This pathway involves the decarboxylation of l -malic acid to l -lactic acid and CO 2. The CO 2 produced by MLF is partially used by carbonic anhydrase to form bicarbonate, further increasing the cytosolic pH.

MLF has also a protective role for S. Under acid stress conditions, lactococci and thermophilic lactobacilli metabolize citrate, producing pyruvate. Citrate metabolism requires the following three steps: i citrate intake via permease CitP , ii formation of oxaloacetate from pyruvate through the activity of citrate lyase CL , and iii decarboxylation of oxaloacetate into pyruvate via oxaloacetate decarboxylase OAD.

The end products are acetic acid, CO 2 , and pyruvate. As described above, the environmental conditions determine the fate of pyruvate. Because of the increased pH in the cytoplasm and extracellularly, LAB take advantage of the cofermentation of citrate with other carbohydrates, such as lactose or glucose.

The main protective effects of cofermentation differ among LAB. In the presence of glucose and citrate, homofermentative and facultatively heterofermentative species e. Each mole of citrate produces pyruvate and, subsequently, acetoin, butanediol, and diacetyl, which protect the cell against acid stress. Furthermore, Lc. One additional mole each of acetic acid and ATP is synthetized for each mole of citrate metabolized. The metabolism of nitrogen, which includes proteolytic and urease systems, plays a pivotal role in LAB growth and survival.

The regulation of nitrogen metabolism occurs mainly via GlnR and CodY , Under conditions of high nitrogen availability, GlnR controls the import of nitrogen-containing compounds and the synthesis of intracellular NH 3. On the other hand, CodY interacts with its BCAA coeffector to control amino acid biosynthesis, transport systems, and peptidases. Genomes of LAB encode extracellular or cell wall-associated proteinases that hydrolyze environmental proteins into oligopeptides, as well as specific transport systems to take up oligo-, di-, and tripeptides and FAA from the environment.

Intracellular oligopeptidases and peptidases hydrolyze oligopeptides and peptides into smaller peptides and FAA. LAB modify proteolysis as a response to environmental stresses. Under acidic conditions, they reduce the abundance of amino acid transporters, and FAA become limiting.

Consequently, many lactobacilli increase the activities of peptidases, such as PepN and PepO , , , The proteolytic system of LAB is also influenced by carbon metabolism VBNC cells of Lc. Much more needs to be uncovered regarding the mechanisms of carbon and nitrogen coregulation. CcpA forms complexes with CodY in B.

Recently, it was shown that the aminotransferase IlvE of S. The metabolism of FAA not only contributes to flavor development in fermented foods but also is crucial for LAB stress resistance. It allows the production of precursors for the biosynthesis of amino acids, fatty acids, nucleotides, and vitamins, generates energy under starvation conditions, and increases the intracellular pH Fig. Deamination and decarboxylation of FAA are the main routes that enhance LAB growth and survival under environmental stress conditions.

Amino acid decarboxylation leads to the conversion of an amino acid into the corresponding amine and CO 2. This reaction is coupled to the exchange between substrate and product e. The translocation is driven by the opposite gradients of substrate and product over the membrane, which are maintained by decarboxylation.

These combined activities generate a PMF because proton consumption during decarboxylation leads to an increase of the intracellular pH Several LAB genomes e. The main FAA pathways induced under environmental stress responses are discussed in the following paragraphs.

All genetic components of this pathway are present in LAB genomes Nutrient sources e. The ADI pathway of Streptococcus rattus is subject to substrate induction and catabolite repression The NH 3 produced alkalizes the cytoplasm, thus increasing acid resistance , In addition, the extra ATP produced via the ADI pathway enhances cell survival after carbohydrate depletion , while the intermediate carbamoyl phosphate is used to synthesize pyrimidines. The intergenic regions of the arcABC genes have several cis -regulatory element sites, which is suggestive of direct regulation of the ADI pathway through CcpA , The arcABC genes of Lc.

This pathway allows the hydrolysis of agmatine into putrescine, NH 3 , CO 2 , and ATP and is a response mechanism to acid and starvation stresses. It represents a potential health risk for humans due to the synthesis of putrescine, a biogenic amine. The AgDI pathway of S. GAD catalyzes the decarboxylation of glutamate into GABA, a bioactive compound that acts as a neurotransmitter in humans.

The uptake of glutamate and the expulsion of GABA are mediated by the electrogenic antiporter, which, together with the decarboxylation of glutamate, produces PMF. Three decarboxylation-antiporter cycles are needed to synthesize 1 mol of ATP. Concomitantly, the extracellular pH increases slightly due to the exchange between glutamate and the more alkaline compound GABA. The uptake of aspartic acid and the expulsion of alanine are mediated by an electrogenic antiporter, which, together with the decarboxylation reaction, produces PMF This pathway has been described for Lactobacillus sp.

M3 , Lactobacillus buchneri , and Lb. Other LAB, including Lb. The exact mechanism of regulation of the AspD pathway is presently unknown. In these organisms, hdcA forms an operon with a downstream gene hdcB of unknown function. HDC activity is regulated via the cytosolic pH. The enzyme is active at low pH, but a neutral or alkaline pH disrupts its substrate-binding site. The survival of Lc. BcaT and aromatic amino acid aminotransferases AraT play a pivotal role in the regulation of the proteolytic system of Lc.

BcaT is repressed by glucose, which confirms that a link exists between carbon and nitrogen regulation. The hydrolysis of urea enhances survival of LAB under acid stress conditions. Among LAB, the urease system has been studied mainly in S. The urease system produces NH 3 and CO 2 from urea, thus protecting cells against acid stress. CO 2 serves in several anaplerotic reactions that are responsible for the biosynthesis of amino acids and nucleic acids.

The urease operon of the oral LAB S. The urease system is induced when S. The expression of the urease operon in S. I is mediated via CodY Inorganic polyphosphate poly-P is a polymer of phosphoanhydride-linked phosphate residues that can be found as chains of a few to thousands of residues in living organisms In bacteria, the accumulation of poly-P is dependent on the activity of polyphosphate kinase Ppk; EC 2. The hydrolysis of poly-P is catalyzed by exopolyphosphatases Ppx; EC 3.

Lactobacillus organisms, such as Lb. The ability of lactobacilli to accumulate poly-P is correlated with the presence of ppk genes in the strains and the availability of phosphate in the medium , However, Lb. Poly-P has a protective role against oxidative stress in bacteria, where it acts as a chaperone preventing the aggregation of damaged proteins The synthesis of poly-P in Escherichia coli is regulated by hypochlorous acid Poly-P is also involved in the regulation of general stress resistance pathways in different bacteria , but more research is required to fully understand its role in LAB Some stresses are imposed on cells through specific molecules, such as protons, ROS, metal ions, and salts such as NaCl.

LAB have developed various mechanisms to directly counteract the negative effects of these molecules Fig. Consequently, toxic molecules are not always associated with a single type of stress. Schematic representation of the main molecular mechanisms preventing macromolecules from being damaged in LAB. SOD, superoxide dismutase. Among the many mechanisms that LAB species employ to counteract stress damage, the F-ATPase system deserves attention as a system that protects against protons.

The F-ATPase acts in parallel to the above-described metabolic adaptations that result in alkalization of the intracellular and extracellular environments. F-ATPase is a reversible enzyme that can also synthesize ATP by using protons that flow from the environment into the cell. This reaction requires ATP because the expulsion of protons from the cytoplasm to the outside environment takes place against an increasing proton gradient that is a consequence of the synthesis of lactic acid during carbohydrate fermentation Distinctly aciduric oral streptococci, such as S.

The capacity to pump protons out of the cell under acidic conditions protects important glycolytic enzymes and allows long-term survival Lactobacilli increase the synthesis of one or more of their ATP synthase components e. The F-ATPase complex not only is involved in the maintenance of cytoplasmic pH homeostasis but also responds to bile salt exposure , — It was noticed that in Lb. Oxygen per se does not damage cells, but during the operation of metabolic pathways it is partially reduced to water, leading to the synthesis of ROS, including superoxide anion radicals, hydroxyl radicals, and H 2 O 2.

These display a considerable oxidizing potential and are highly toxic to cells , As detailed above, Lc. Superoxide anion as well as hydroxyl radicals are produced during the Fenton reaction , Metal ions can catalyze certain reactions leading to the formation of ROS, which is in fact considered one of the underlying mechanisms of metal-induced stress.

Iron, copper, and other redox-active metal ions exert their effects by stimulating the Fenton reaction. At the molecular level, ROS can react with macromolecules, such as proteins, lipids, and nucleic acids. Superoxide anions have a limited oxidizing potential and may attack compounds such as polyphenols, ascorbate, and catecholamines The damage caused by the hydroxyl radical, which is the most destructive oxidative agent, includes strand breakage and chemical modification of DNA , Bacteria such as Lc.

Notably, induction of genes encoding ROS-protective enzymes is growth phase dependent and confers multistress resistance , NADH peroxidase activity is rather low in Lc. It was recently proposed that metabolically synthetized pyruvate reacts nonenzymatically with residual H 2 O 2 to produce water They catalyze either the two-electron reduction of oxygen to H 2 O 2 performed by Nox1 or the four-electron reduction of oxygen to water AhpCF and thereby provide a very efficient enzymatic defense against oxidative stress Another key player in resistance to oxidative stress is SOD, which eliminates superoxide anion radicals.

Bacteria possess different types of SOD enzymes depending on the bivalent metal cofactor with which the enzyme is coupled, i. Analysis of an Lc. An alternative strategy to counteract superoxide anion radicals is by the production of high levels of intracellular glutathione When this pathway is impaired by restricting access to these nutrients, extracellular superoxide is generated by partially reduced demethylmenaquinones upon exposure to oxygen Thus, E. Bacteria can also cope with oxidative stress through catalase, an enzyme catalyzing the detoxification of H 2 O 2.

The following three classes of bacterial catalases are currently recognized: monofunctional catalases, catalase-peroxidases, and manganese catalases pseudocatalases The monofunctional catalases and the catalase-peroxidases contain heme as a prosthetic group. The monofunctional catalase specified by the genome of E.

In order to retain the low intracellular redox potential required to keep proteins in their reduced form, the presence of glutathione as well as the thioredoxin systems is crucial Members of the genus Streptococcus contain glutathione. They either synthesize glutathione or import it from the medium In addition, streptococci encode a glutathione reductase, which catalyzes the reduction of glutathione disulfide that is produced upon oxidation of glutathione by NADPH The glutathione reductase gene in S.

This relatively large genetic repertoire emphasizes the importance of such enzymes in oxidative stress resistance for a review, see reference Adaptation is also part of the survival strategy of LAB to cope with oxidative stress. Upon exposure to sublethal concentrations of H 2 O 2 , Lc. PoxB utilizes oxygen to convert pyruvate to acetyl-phosphate Fig. Detoxification of the latter molecule is subsequently done by NADH peroxidase Notably, PoxB activity is induced by oxygen or H 2 O 2 and repressed by glucose In silico analyses of Lb.

This is suggestive of regulation of poxB transcription in response to oxidative stress. Other specific mechanisms involved in the adaptation of S. Genome analysis of S. The mechanisms that evolved in LAB to overcome oxidative stress seem to overlap, and thus cells subjected to a specific stress condition can trigger different stress responses for a review, see reference The cAMP receptor protein, the fumarate and nitrate reduction regulator FNR , and FNR-like proteins Flp are part of a superfamily of proteins that encompasses structurally related transcription factors implicated in the control of stress responses As mentioned above, such proteins display structural features allowing transmission of environmental and metabolic signals to the cell and which may trigger general and specific responses to various stressful physiological conditions In Lb.

They are located in two paralogous operons in which the proximal genes are predicted to encode metal ion transport systems It was postulated that Lc. Many regulators in streptococci modulate the interplay between oxidative stress and iron homeostasis, virulence, and competence Iron is considered a key element in regulatory proteins sensing redox modifications in the presence of oxygen This is illustrated by an S.

In fact, control of iron homeostasis is considered to be coupled to ROS protection in streptococci Also, the S. Its involvement in the regulation of proteins related to oxidative stress was revealed when the rgg gene was inactivated Inactivation of an Rgg-like protein of S. Osmotically active compounds serve a dual function in osmoregulation. In addition to maintaining a stable cell turgor, they also exert protective effects on biomolecules in vitro during stressful challenges.

Various osmotically active compounds, such as proline and betaine, protect soluble and peripherally membrane-bound proteins from the denaturing effects of modifications in intracellular ionic strength and water availability. Osmotically active compounds can interact with proteins in several different ways, such as preferential exclusion, water replacement, hydration force, and vitrification of sugars Betaine transport activity in LAB, such as Lc.

Other osmotically active compounds, such as the disaccharide sucrose, protect Lc. The protective effect of ionic solutes has been ascribed to the intracellular accumulation of compatible solutes as a response to osmotic stress. Thus, ionic solutes e. A large number of bacterial ABC-type drug export systems have been characterized Most of these microbial drug extrusion systems are specific chemical or multidrug resistance MDR systems that remove various substances from the cytoplasm or membrane of a cell Notably, LmrA expressed in human or insect cells efficiently exports drugs out of these cells and complements a lack of expression of the human MDR1 P-glycoprotein A homolog of lmrA with highly significant structural and functional similarities to its lactococcal equivalent has been identified in O.

In bacteria, as in all living cells, the cellular proteome is in a constant state of flux. Protein quality control, including refolding or degradation of damaged proteins, plays an indispensable role under both stressed and nonstressed conditions. Similarly, the catalog of cellular proteins is continuously being modified to meet the challenges arising from changes in the environment.

The synthesis of chaperones and proteases is quickly induced under stress conditions to combat the potentially deleterious aggregation of denatured proteins Fig. While the task of chaperones is to protect functional proteins and to refold misfolded ones, proteases provide the last line of defense by removing irreversibly damaged proteins, after which the amino acids are recycled.

Under changing conditions, proteolysis also removes functional proteins for regulatory purposes or simply because they are no longer needed. It is obvious that the balance between refolding and proteolysis, including the highly specific recognition of substrates, is crucial to avoid wasteful destruction , Schematic representation of the main molecular mechanisms repairing damaged macromolecules in LAB.

Chaperone proteins recognize stretches of hydrophobic amino acids typically exposed by nonnative proteins and bind to these regions to prevent aggregation. They can also actively assist protein folding in an ATP-dependent manner. One well-conserved bacterial chaperone system that can efficiently refold misfolded proteins is that of DnaK Hsp A deletion in dnaK resulted in a thermosensitive phenotype for Lc. DnaK is essential in S.

The inability of DnaK proteins of S. Since only a few reports deal with DnaK structure or biochemical function in LAB, several fundamental questions remain to be clarified. For example, to date, the cochaperone-dependent folding activity of DnaK has not been demonstrated for any LAB. GroES is the most abundant protein in B. The Clp machinery is probably the main system for general protein turnover in LAB, as is the case in other low-GC Gram-positive bacteria The central proteolytic core consists of 14 ClpP subunits stacked back to back in two heptameric rings, forming an internal chamber that encapsulates the active sites of the ClpP peptidases The peptidase multimer associates with one or two hexameric rings of Clp ATPases to gain proteolytic activity.

The Clp ATPases, together with the proteolytic core, are responsible for substrate recognition, unfolding, and translocation into the ClpP degradation chamber. In the absence of ClpP, threading of substrates through the Clp ATPase ring structures leads to disaggregation and refolding of protein substrates, which are activities typical of molecular chaperones ClpE is absent in E. With the exception of clpX , the genes encoding the Clp family of proteins are not essential for most bacteria, including LAB , which has facilitated the construction of knockout mutants and the study of the physiological roles of Clp proteins in a variety of LAB species.

In contrast, ClpX is not essential in S. Phenotypic studies with mutant strains lacking functional ClpP, ClpL, ClpE, or ClpB revealed the central role of these proteins under stress conditions and, for pathogenic LAB species, in virulence , , — In contrast to the situation in B.

Phenotypic characterization of an S. The alternative sigma factor ComX, which drives the expression of competence genes, was shown in S. This task is performed by ClpEP in S. The primary stress induction of molecular chaperones and Clp proteins in LAB occurs at the level of transcription and is regulated mainly by the transcriptional repressors CtsR and HrcA. The B. One of the repressors is absent in some LAB species, and a single regulator controls expression of both regulons The expression of S.

The HrcA and CtsR regulons may partially overlap in several other members of the Streptococcaceae , Sequence analysis of Lb. An example of reductive evolution of stress repressors with a different outcome is provided by Lb. The HrcA regulon in these species has expanded to include clp genes, in addition to genes encoding the groEL and dnaK operons. According to studies of other organisms, HrcA is prone to aggregation, a characteristic feature that is also central to its autoregulation HrcA is thought to be released from ribosomes in an inactive form.

Thus, protein stress directly affects HrcA activity. Molecular studies have revealed several differences in the mechanisms of regulation of CtsR function between LAB and other bacteria. McsB is not involved in the regulation of CtsR activity during heat stress in vivo , and CtsR itself acts as an intrinsic thermosensor whose DNA-binding activity gradually decreases with increasing temperature. The CtsR regulon is also derepressed under several environmental stress conditions other than heat shock, such as acid stress and oxidative damage.

This implies that alternative mechanisms for CtsR inactivation must exist. A recent report provided evidence that S. The FtsH proteins of Lc. Inactivation of ftsH resulted in salt-, heat-, and cold-sensitive phenotypes of Lc. The mechanisms controlling ftsH expression in other LAB have not been studied. HtrA high temperature requirement protein A , also known as DegP, is involved in degradation of extracellular polypeptides and misfolded proteins Fig. HtrA contains a chaperone and a proteolytic domain.

An Lc. The htrA gene has been identified as a heat shock gene in Lactobacillus helveticus , and elevated HtrA levels have been detected in Lb. However, the regulatory mechanism behind the stress-inducible expression of htrA in LAB remains to be elucidated. The members of the sHSP family function as folding chaperones, assisting the protein folding process by stabilizing unfolded or partially folded proteins without actively promoting their remodeling Furthermore, sHSPs appear to play an important role in maintaining membrane integrity, especially under stress conditions Lo18 has been suggested to play a dual role in O.

A sudden temperature downshift induces a cold shock response, during which growth of cells is first arrested for a period before reinitiation at a new and much reduced rate. The cellular effects of cold shock include the following: i a decrease in membrane fluidity, ii a reduced efficiency of mRNA translation and transcription through stabilization of secondary structures of DNA and RNA, iii ineffective or slow protein folding, and iv hindered ribosome function In cold-shocked E.

Members of the CSP family enhance the expression of cold shock-inducible genes by destabilizing nonproductive secondary structures in mRNA at low temperatures and therefore are often called RNA chaperones While cold and heat shocks are considered highly separate processes in E. Cold shock of S. Furthermore, the clpL knockout mutant of S.

Transcriptional analyses have indicated that clpX is induced by both heat and cold in Lc. The proposed cross talk between cold and heat shock responses in LAB is further supported by the improved cryotolerance seen after heat treatment of Lb. Similar findings may link acid and cold stress responses in LAB; for Lb. Spx is a global transcriptional regulator of oxidative stress in several Gram-positive bacteria and plays important roles as both a positive and a negative modulator of gene expression Studies of B.

DNA microarray work with spx mutants confirmed the function of Spx as a major oxidative stress regulator , Streptococci harbor two bona fide Spx regulators, whereas enterococci have only one , — According to the current DNA damage paradigm, cells undergo an inducible process called the SOS response during genotoxic stress. This response triggers the expression of genes involved in DNA replication, repair, and mutagenesis The SOS response is repressed under physiological conditions by binding of the LexA repressor protein to operator sites in the promoter regions of SOS genes, including its own gene, lexA While the bacterial DNA damage response is one of the most extensively studied stress responses, it has not been studied widely in LAB.

The uvrA gene, which encodes a nucleotide excision repair protein and is a typical member of LexA regulons, has been characterized for S. Other DNA repair activities that have been implicated in the resistance of S.

LAB have cell envelopes that are typical of Gram-positive bacteria. The cell wall itself comprises a PG sacculus that is usually decorated with lipo teichoic acids, polysaccharides, and proteins for a review, see reference The cell wall provides a point of non covalent attachment of important extracellular enzymes and external structures, such as pili and flagella It also participates in processes of adhesion, cell aggregation, and cell division Most importantly for this review, the cell envelope acts as a diffusion barrier between the cell and its surroundings.

It is crucial for maintaining the integrity and shape of the cell and for resisting changes in osmotic pressure. Many of the environmental or technological stressors that LAB can encounter have an impact on the cell envelope. This is why it is very difficult to arrive at a strict definition of cell envelope stress, as regulator systems that respond to stresses exerted on or sensed at the cell envelope often also play a role in the normal physiology of the cell The cell wall is a major target in our fight against pathogenic bacteria, including pathogenic LAB.

Cell wall synthesis can be inhibited by antibiotics that block certain enzymatic steps or capture one of the precursors Antibiotics such as vancomycin, lantibiotics, or bacitracin target the translocation via lipid II of PG precursors across the cytoplasmic membrane lipid II cycle Vancomycin binds to the d -alanyl— d -alanine ends of PG precursors, thereby inhibiting the formation of mature PG Inducible vancomycin resistance was first described for clinical isolates of enterococci , So far, eight enterococcal van gene clusters vanA to - E , vanG , vanL , and vanM have been described, and they encode ligases required for the synthesis of either d -Ala— d -lactate or d -Ala— d -Ser PG precursors for which glycopeptides, such as vancomycin, have low affinity , The molecular mechanisms of regulation of vancomycin resistance gene expression have been reviewed thoroughly and are not discussed here.

The specific ligand recognized by the VanS sensor proteins has not yet been determined unequivocally. Two distinct models have been proposed: direct induction, in which the sensor kinase binds the antibiotic, and indirect induction, in which the sensor kinase binds a cell wall component that either is an intermediate of cell wall biosynthesis or accumulates as a result of antibiotic action Lantibiotics, such as nisin produced by Lc.

Nisin binds lipid II and thus has a double effect on the cell: it inhibits PG synthesis, and it forms pores in the cytoplasmic membrane through which essential components can leak out, killing the cell The main resistance mechanism against positively charged molecules, such as lantibiotics, is a change of the overall negative charge of the cell wall by coupling the positively charged amino acid d -alanine to teichoic acids in the cell wall Bacitracin, a cyclic dodecylpeptide antibiotic that blocks the lipid II cycle by inhibiting dephosphorylation of isoprenyl pyrophosphate , is counteracted by the action of specific ABC transporters BceAB see below.

Other mechanisms, such as the synthesis of alternative isoprenyl pyrophosphate phosphatases or de novo synthesis of undecaprenyl phosphate, have also been described , BceRS was first described for B. A remarkable characteristic of these systems is that IM-HKs are unable to function as sensors: this function is provided by the ABC transporter, with which they are usually genetically and functionally associated , These studies revealed remarkable differences in the genetic organization and regulons.

Both TCSs constitute functional units with their corresponding ABC transporters, and transcriptional analyses showed that while TCS09 controlled the expression of the cognate ABC09 transporter, TCS12 controlled the expression of the orphan ABC as well as the dlt operon involved in d -alanylation of teichoic acids and the mprF gene involved in l -lysinylation of phospholipids One of the ABC transporters is essential for sensing bacitracin and eliciting a response A later study identified four additional genes that belong to the MbrABRS regulon, including those for the cell surface antigen SpaP and for MurM and MurN, which are alanine-adding enzymes involved in the branched-peptide PG biosynthetic pathway.

Although most OCSs are predicted to be cytosolic proteins, some may actually be membrane bound The BcrR bacitracin sensor of E. An auxiliary protein is not required for bacitracin sensing by BcrR Deletion of the transmembrane domain abolished DNA-binding activity, suggesting that membrane localization is essential for BcrR activity Binding of bacitracin did not alter the DNase I protection pattern or the apparent DNA-binding constant for binding of BcrR in vitro to the bcrABD promoter, but it did lead to induction of transcription of this promoter in an in vitro transcription assay On the basis of these results, BcrR is proposed to always be bound to its target DNA sequence; binding of bacitracin would induce a conformational change facilitating a productive interaction with the RNA polymerase, through an as yet undetermined mechanism One of the main functions of the cell wall is to maintain cell shape and to counteract the high internal osmotic pressure, which can reach up to 2, kPa in Gram-positive bacteria.

The cell envelope plays a key role in the regulation of osmotic stress, as it is highly permeable to water, while the cytoplasmic membrane acts as a barrier for most solutes. The main response of LAB to osmotic stress is the intracellular accumulation of osmoprotectants, of which the most common are glycine betaine, choline, carnitine, and dimethylsulfonioacetate Due to their reduced biosynthetic capabilities, LAB have to import these compounds from their natural environments, such as milk, meat products, or plant materials , The ability of these osmolytes to reduce the inhibitory effect of hyperosmotic situations has been established for a large number of LAB , — Another mechanism for LAB to cope with osmotic stress is through regulation of the intracellular concentrations of certain amino acids.

Intracellular concentrations of proline, glutamate, and sometimes alanine and aspartate increase when Lb. Some amino acids reduced the impact of osmotic stress on growth when they were added exogenously to LAB cultured in chemically defined media. This was reported for the addition of proline to Lc. Di- and tripeptides generated by the activity of cell envelope-associated proteinases can also act as osmoprotectants in LAB, as reported for O.

LAB species have developed efficient mechanisms for recruitment of osmoprotectants from their environment. Accumulation of osmolytes is mediated by transporters that move solutes across the membrane against their electrochemical gradients in processes that require metabolic energy Interaction of the transporters with membrane lipids determines their activity in response to osmotic changes Betaine and proline uptake in Lc. This operon encodes a transport system of the ABC family that uses two ATP molecules per translocated molecule of glycine betaine OpuA detects changes in the intracellular ionic strength, and its activity depends on electrostatic interactions between lipid head groups and OpuA In particular, sensing depends on the C-terminal tail of the CBS domains, which is negatively charged A model was proposed in which the transporter switches from the active to the inactive state depending on the interaction of the CBS domains with lipid head groups in the membrane.

Repulsion between the charged C-terminal tail and the anionic heads of the membrane lipids counteracts the interaction between the CBS domains and the membrane. In this way, the intracellular ionic strength and the concentration of anionic lipids in the cytoplasmic membrane determine the activation of the transporter A later study showed that the interaction of the CBS domains and the membrane lipids depends on a surface-exposed cationic region in the CBS domain, in agreement with the proposed model A genome-wide gene transcription analysis revealed that Lc.

In Lc. The uptake of osmolytes by Lb. This system has a high affinity for glycine betaine and a low affinity for proline and is expressed semiconstitutively, i. When bacteria are faced with a sudden drop of the external osmotic pressure, they use mechanosensitive channels MSC to counteract the effects. The MSC act as emergency release valves, allowing the expulsion of osmolytes and water.

MSC share an organization of a C-terminal cytoplasmic domain and an N-terminal transmembrane domain formed by symmetrical helices that can change between open and closed states Although their mechanism of sensing is still unclear, it is well known that MSC can sense changes in the tension of the lipid bilayer of the cytoplasmic membrane Most microbes appear to possess members of one or both families of bacterial MSC Although Lc.

LAB can counteract several stresses by modifying their cytoplasmic membranes. The regulation of membrane fluidity in bacteria is a rather complex process that often involves universal protein chaperones, such as GroEL and sHSPs , The fluidity of the cytoplasmic membrane is highly influenced by the ratio of unsaturated to saturated fatty acids, and alterations in membrane fatty acids may represent a generalized response utilized by many bacteria to survive environmental stresses affecting the membrane.

The ratios of unsaturated to saturated and cyclic to saturated membrane fatty acids decreased in a strain of Lb. The decrease in membrane fluidity in Lactobacillus could be compensated by other variations, such as an increase of cyclic fatty acids Applying a cold, acid, or ethanol shock to O. When Lb. High salt concentrations led to an increase of the size of the cells and to sensitivity to antimicrobial peptides targeting PG, such as nisin These effects were associated with a reduction of the cross-linking of PG molecules due to altered expression and activity of PBPs , Osmotic stress induced by a high salt concentration in Lb.

Exposure of Lc. This system also responds to the presence of other antibiotics that interfere with the lipid II cycle vancomycin and ramoplanin , to the action of detergents and organic solvents, and to other unspecific stimuli on the cell envelope , — Disrupting the gene for CesR led to a reduction in the ability to resist salt and osmotic stresses The authors could not generate a CesS mutant, probably because that mutation was lethal.

Nevertheless, a nisin-resistant strain obtained through a nisin adaptation process overexpressed cesFSR and its regulon The cesFSR system is also induced by vancomycin, bacitracin, and the bacteriocins plantaricin C and Lcn Analysis of the transcriptome of Lc. Many of the genes in the cesFSR regulon of Lc.

The cooverexpression of these genes increased the resistance of Lc. Growth of Lc. A bp conserved motif essential for LiaR binding is present upstream of the genes directly controlled by the LiaFSR system Disruption of this system in S. Deletion of the LiaR homolog in E. This deletion also restored susceptibility to daptomycin and telavancin, antibiotics that disrupt the cell membrane, in a multidrug-resistant E.

All in all, the LiaFSR system homologs have a widely conserved cell envelope stress-sensing function in Firmicutes , although the regulons differ quite considerably among genera. The regulons in LAB are broader than that in B. LAB employed for potentially health-promoting applications are mainly lactobacilli. They encounter a variety of stresses during production and use.

Before a probiotic bacterium begins to fulfill its biological role in the gut, it must survive a battery of environmental challenges Initially, probiotic bacteria must be processed in a suitable product form to enable oral consumption in adequate numbers, and then the bacteria must survive the harsh conditions imposed during passage through the GIT Although they are not strictly considered probiotics, increasing evidence has shown that dead bacteria can still have a health-promoting effect Probiotics are normally grown to high numbers before undergoing a drying process to produce a high-cell-density dried powder formulation.

Such extreme temperatures can affect membrane fluidity and compromise cellular integrity and basic cell processes, such as ribosomal function, protein folding, and enzymatic activity Some species of lactobacilli are relatively thermotolerant; for example, Lb. The effect of heat shock and the induction of a stress response have been studied in a variety of lactobacilli see above.

In general, the resistance to stress is higher when cells are previously exposed and adapted to a sublethal treatment with a homologous stress. For example, enhanced heat tolerance was obtained when Lb. Similarly, treatment of Lb. This improvement was associated with induction of the chaperone GroEL. Heat stress was previously shown to enhance the expression of the heat shock genes hsp As mentioned earlier, CSPs are induced as a result of cold shock pretreatment, and they have been associated with the stabilization of mRNA Three induced CSPs were previously identified in Lb.

In addition to temperature stress, spray-drying also exposes cells to osmotic stress, dehydration, and oxidative stress A sudden increase in the environmental osmolarity hyperosmotic stress results in the movement of water from the cell to the outside, causing a detrimental loss of cell turgor pressure and changing the intracellular solute concentration and cell volume, which ultimately can seriously affect cell viability Probiotic bacteria can develop significant physiological changes to respond to osmotic stress conditions.

Expression of the htrA gene increased 8-fold in Lb. Probiotic LAB can also adapt to hyperosmotic conditions by accumulating compatible solutes, such as betaine, carnitine, and trehalose , , For example, Lb. The same was found for Lb.

In a study on Lb. This change was explained in terms of an increased activity of the oxygen-consuming fatty acid desaturase system, which serves to reduce free radical damage in the cell. The principal stresses encountered by probiotic bacteria during passage through the stomach and upper intestinal tract include the low pH in the stomach as well as the detergent-like properties of bile in the duodenum.

Exposure to these stresses causes damage to the cell envelope, DNA, and proteins The survival of probiotic lactobacilli in low-pH environments can be increased hugely several orders of magnitude in the presence of metabolizable sugars, such as glucose Other systems to regulate internal pH also exist; for example, Lb.

Similarly, amino acid decarboxylation combined with an amino acid antiporter leads to the biochemical consumption of protons, thus contributing to acid tolerance during growth , The exposure of Lb. As detailed above, the Clp chaperones actively promote refolding or degradation of damaged proteins Acid stress can also cause molecular changes in the cell surface.

Moreover, bacteria are able to change their membrane lipid composition in response to external stresses. The composition of the cytoplasmic membrane can have a dramatic effect on the ability of probiotic bacteria to survive stresses ranging from heat to low pH.

For example, the membranes of acid-adapted cells of Lb. In the latter strain, the membrane fatty acid composition and in particular the saturation level is associated with changes in cell survival during heat or acid shock. Probiotic LAB can also sense and develop resistance mechanisms against bile salts. For example, an eight-gene operon encoding a TCS, a transporter, an oxidoreductase, and four hypothetical proteins has been implicated in sensing bile salt in Lb.

Likewise, the genes encoding multidrug transporters driving extrusion of bile salts are upregulated in Lb. Indeed, mutations in the lr gene encoding an MDR transporter impaired the ability of Lb. Exposure of Lb. This is in agreement with the observation that mutations in lr clpL in Lb. Some probiotic LAB can efficiently express a range of bile salt hydrolases BSH that confer protection against bile via bile salt deconjugation The cellular response toward bile of the well-documented probiotic Lb.

Production of external EPS layers can have a significant protective effect against bile and other harsh environmental conditions. Indeed, strains producing EPS exhibit significantly more growth in the presence of 0. Although intrinsic bile tolerance appears to be strain dependent, probiotic strains can be made to adapt progressively to the presence of bile salts by subculture in gradually increasing concentrations of bile In considering probiotics, it is also important that stationary-phase cells are generally more resistant than log-phase cells to various stressors, since many stress resistance mechanisms are switched on when active growth ceases.

This is an important consideration, since probiotic LAB are generally prepared from stationary-phase cultures. Some of the approaches aim at improving culture viability under the conditions of spray-drying, a procedure increasingly employed to concentrate and preserve particular probiotic cultures used in functional foods and health supplements. Thus, supplementation of the growth medium with saturated fatty acids is likely to increase the acid and heat tolerance. Presumably, this effect is due to increased rigidity of the cytoplasmic membrane.

Similarly, overexpression of HSPs in Lb. Interestingly, heterologous expression of the BilE transporter in Lc. Other authors have emphasized the importance of preconditioning cells with homologous stressors, particularly heat shock prior to spray-drying Bile-adapted strains usually display cross-resistance to a variety of other stress factors. Finally, culture reconstitution conditions should be optimized for each probiotic strain in order to achieve accurate viable probiotic numbers from dried cultures To colonize and thrive within the mammalian host, bacteria must cope with a variety of environmental insults.

These include fluctuations in temperature, pH, and nutrient source and availability, the presence of ROS, reactive nitrogen species RNS , and antimicrobial peptides generated by professional phagocytic immune cells, and, eventually, the different types of stresses brought about by administered antibiotics.

Given that the ability to sense and respond to environmental stresses is an essential survival trait in bacteria, it is not surprising that stress responses are often intertwined with virulence. In the case of pathogenic LAB, which belong mainly to species within the Streptococcus and Enterococcus genera, multiple examples of stress survival pathways that participate in virulence have emerged in the past 2 decades.

They do so directly, by facilitating bacterial survival and dissemination, or indirectly, by controlling the expression of virulence factors. This section focuses on important paradigms of stress regulators as well as stress-adaptive mechanisms that have been linked to virulence in streptococci and enterococci Table 1.

As already discussed above, general stress responses are triggered by different stress conditions that result in the accumulation of misfolded proteins, inducing DnaK, GroEL, ClpP, and HtrA. The genes encoding the ClpP peptidase and the Clp ATPase partners are by far the best-studied streptococcal stress genes in terms of virulence potential. The virulence of S. The core genome of S. For an S. Transcriptomic and proteomic analyses indicated that ClpE affects pneumococcal pathogenesis by modulating the expression of important virulence determinants and metabolism-related factors Despite participating in several important physiological processes , , ClpC is apparently not required for pneumococcal virulence , , while inactivation of clpX is lethal Virulence of a mutant GBS, S.

The involvement of Clp ATPases has not been studied extensively in enterococci. Deletion of clpB reduced the ability of E.

Ваша мысль how to read betting lines mlb schedule блестящая

In a note to his fellow Republican senators just before the House was to begin voting, he said he is undecided. A majority of the U. The House vote on an article of impeachment for incitement of insurrection was still underway Wednesday, but the Democratic-led House had secured enough votes to impeach Trump. Some Republicans joined Democrats in voting to impeach the president.

Five people died. It says Biden received a briefing Wednesday on preparations for the event from top law enforcement officials. Capitol, and a number of protests are planned both in Washington and in state capitols and cities across the U. Newly elected Michigan Rep. Meijer announced he would vote to impeach Trump in a statement released Wednesday as the House was debating the proceedings.

Meijer said impeaching Trump will likely exacerbate division and set precedent. Voting is underway in the House on impeaching President Donald Trump over the violent siege at the U. Capitol last week by a mob of his supporters. Lawmakers are voting Wednesday on impeaching Trump on a single charge, incitement of insurrection. If it passes, Trump would be the first president to be impeached twice.

The riot has forced a reckoning among some Republicans, who have stood by Trump throughout his presidency and largely allowed him to spread false attacks against the integrity of the election. Trump has taken no responsibility for the riot, suggesting it was the drive to oust him rather than his actions around the bloody riot that was dividing the country.

Democratic Rep. Pressley said in a statement that Conan Harris received a positive test Tuesday evening. Pressley says she received a negative test result. Harris had accompanied Pressley to her swearing-in ceremony last week and was with her in the Capitol complex throughout the Jan.

Harris has shown mild symptoms and remains in isolation, along with Pressley and staff who were in close contact. Senate Majority Leader Mitch McConnell has said in a note to his fellow Republican senators that he is undecided on whether President Donald Trump should be convicted if the House votes to impeach him.

The House is poised to vote to impeach Trump on Wednesday for a second time after he egged on a violent mob of his supporters who invaded the Capitol last week. President Donald Trump says he opposes violence in a statement read on the House floor as members debate impeaching him for his role in fomenting the violent insurrection at the Capitol last week. I call on ALL Americans to help ease tensions and calm tempers.

Five people died, including a U. Capitol Police officer. The president falsely claimed widespread voter fraud cost him the election won by Democrat Joe Biden. Capitol has exceeded The count by The Associated Press resulted from a nationwide review of court records and announcements of arrests issued by law enforcement agencies.

The charges range from misdemeanour curfew violations in the District of Columbia to federal felonies related to the assault of law enforcement officers, theft of government property and possessing firearms and explosives. Federal prosecutors and the FBI said this week they are pursing dozens more suspects who have been identified through photos and videos from the Jan.

Newhouse of Washington has added his name to the short list of Republicans supporting the impeachment of President Donald Trump. He said Tuesday on the House floor that the article of impeachment is flawed, but he will not use process as an excuse to vote no. Newhouse says the president took an oath to defend the Constitution against all enemies foreign and domestic.

But McCarthy also says he believes it would be a mistake to impeach Trump in such a short time frame. Trump leaves office on Jan. The House is set to vote later Wednesday on impeaching Trump, accusing him of rallying the violent mob. The aide was not authorized to discuss the matter publicly and spoke on condition of anonynmity. Michael A. Bolton is the inspector general and he will lead the review. It will focus on security preparations for the Jan. Capitol Police Chief Steven Sund resigned in the wake of the riot, as did the top security officials in the House and Senate.

The department is facing intense scrutiny after its lacklustre response to the riot, poor planning and failure to anticipate the seriousness of the threat drew widespread condemnation. If the House impeaches President Donald Trump, a Senate trial on whether to convict him of inciting insurrection seems all but certain to have to wait until President-elect Joe Biden is inaugurated. The House is set to vote later Wednesday on impeaching Trump, accusing him of rallying a violent mob of supporters to attack the Capitol last week.

Their campaign on his behalf has raised eyebrows because self-promotion for the prize is considered unseemly. Since then Morocco and Sudan have also agreed to recognize Israel. He must go. He is a clear and present danger to the nation that we all love.? The House is set to vote Wednesday afternoon on impeaching Trump, accusing him of rallying a violent mob of supporters to attack the Capitol last week. Republican Sen. Graham is a frequent ally of President Donald Trump. At least five GOP House members have said they will support impeachment, and two Republican senators have called on Trump to resign.

Another GOP senator has said he will take a look at the articles of impeachment when they are sent to the Senate. The debate is heated almost from the start as the House sets up a vote to impeach President Donald Trump. Democrats and a few Republicans say Trump must be removed immediately after he egged on a violent mob of supporters a week ago who then stormed the Capitol.

Republican Rep. But Democratic Rep. Democratic lawmakers have opened the historic impeachment effort in the House by saying that every moment Donald Trump is in the White House the nation is in danger. Jim McGovern, D-Mass. McGovern says it was Trump and his allies who were stoking the anger of the violent mob. Tom Cole of Oklahoma said Jan. But Cole says the Senate could not even begin to consider impeachment until after President-elect Joe Biden is sworn in.

He says he can think of no action the House can take that would further divide the American people than the actions being taken Wednesday. As the House opens its impeachment hearing, the District of Columbia National Guard says it has been authorized to arm troops assigned to security duty on the U.

Capitol grounds. The Guard said in a statement that the authority was requested by federal authorities and approved by Army Secretary Ryan McCarthy as of approximately 6 p. Up to 15, Guard members are expected to be on duty in coming days in the district to support law enforcement in connection with the Jan.

Authorities are concerned about threats of violence, following the insurrection at the Capitol last week. The House has opened its proceedings Wednesday, poised to impeach President Donald Trump for a second time exactly a week after his supporters stormed the Capitol to protest his election defeat. At least five Republicans have said they will join Democrats in voting to remove Trump from office.

New York Rep. Capitol last week. The Democrat spoke about her experience in an Instagram Live video on Tuesday night. The second-term representative, whose New York district covers part of Queens and the Bronx, had strong condemnation for Trump for inciting the riots, as well as members of his administration who have not invoked the 25th Amendment to remove him from office, and those who voted to overturn the election results.

Adam Kinzinger is predicting more Republicans will join him in voting to impeach President Donald Trump. The House is set to vote Wednesday afternoon on impeaching Trump for a second time, accusing him of rallying a violent mob of supporters to attack the U. Politics Technology Video Weather News. Investigators suspect throttle problem in fatal crash of Sriwijaya Air Flight Tesla's bold move is putting Bitcoin on the mainstream corporate radar.

Djokovic holds off Tiafoe; Venus hobbles out in Australia. None of this actually made any sense. It works now. The dealer neer balked, but the response was incredibaly slow, and they dodn't seem concerned. Antilock system ABS , parking brake, master cylinder, calipers, rotors, pulsation or vibration, squeaking, brake failure or wear.

Caused multiple warning lights. My belief is that there is a problem with the e brake cable and it sticks even though not engaged as I now have to replace brakes and rotters for a second time while only owning the car since Jan of ". The dealer initially didn't believe this was happening, but after several back and forths they agreed to exchange the car for another new one. Seems to be a problem with vw Jetta. Not a problem with my golf.

Just both jettas I owned. We live on a dirt road and it can be very irritating, but since it's not covered under warranty, we cannot afford to have it looked at. USB port doesn't work well". Alternator, starter, hybrid battery and related systems, regular battery, battery cables, engine harness, coil, ignition switch, electronic ignition, distributor or rotor failure, spark plugs and wires failure. I noticed many other VW's on the road that had non functioning tail lights.

Turns out this is a constant problem with this car. Repaired under warranty". Had wiring for horn eaten through twice, engine wires that prevent starting without foot on the brake engine would push button start without foot on the brake.

Had to pay Volkswagen to replace wiring for horn but all they had was the same type of wire coating that is a problem and rodents ate through it a second time. Ended up fixing my self and adding hard plastic overwrap to all wires that I could including engine wires and horn wires. Was disappointed that VW dealer knew this was a problem the first time I had them fix it and they didn't have any additional fixes or alternate wiring without the rodent bait coating.

Took it back and they fixed the problem. Something to do with the computer. VW knows about problem and have not been able to correct it. Plus did not tell me about it wen I bought it. Let me find out 3 months later. Had to go back and drive my Jeep. Weird and buggy? Air-conditioning is anemic and effectiveness worsening over last year. Paint fading, chalking, cracking, or peeling , loose interior and exterior trim or moldings, rust.

So far I've noticed each time and reattached them but the day will come that I will lose the part. What parts: the plug in the bumper, the passenger door reflector a cap in the liner of the trunk. It was delivered with power polisher swirls in the paint and now, five months later, the paint is showing signs of rust lower front passenger door and other places. VW sent me a clay bar to clean the paint. I did not request it. It just arrived the mail. I am going to complain to the dealer and to VW Canada.

I haven't had any accidents so I can't imagine why this would be happening. I'm pretty disappointed because I have a white exterior so it's extremely noticeable. Transmission rebuild or replacement, torque converter, clutch replacement. There had been no slipping or other issues prior to this failure.

Despite the dealer claiming that they couldn't find a defect, the clutch was replaced completely, and Volkswagen paid for most of the repair. Volkswagen replaced the transmission under warranty at miles. Gear selector or linkage, leaks, transmission computer, transmission sensor or solenoid, clutch adjustment, rough shifting, slipping transmission.

Sometimes gears seem to grind and sometimes it's hard to downshift properly. REAL problems sometimes getting into reverse from forward or neutral, or from reverse into forward - and this problem seems to be betting worse in last 6 months. A potential safety hazard, and potential cause of an accident.

Problems shifting from N or 1 into 1 or 2". Muffler, pipes, catalytic converter, exhaust manifold, leaks. Dealership repaired it and it was covered. I think the warning light should be in red in order to identify a problem or potential problem. Accessory belts and pulleys, engine computer, engine mounts, engine knock or ping, oil leaks. Oil leak between the engine and transmission bell housing.

If he hadn't caught it I would have been paying thousands for the repair after warranty. Power or manual windows, locks and latches, tailgate, hatch or trunk, doors or sliding doors, mirrors, seat controls, safety belts, sunroof, convertible top. Cruise control, clock, warning lights, body control module, keyless entry, wiper motor or washer, tire pressure monitor, interior or exterior lights, horn, gauges, 12V power plug, remote engine start, alarm or security system.

Shocks or struts, ball joints, tie rods, wheel bearings, alignment, steering linkage includes rack and pinion , power steering pumps and hoses, leaks , wheel balance, springs or torsion bars, bushings, electronic or air suspension. VW quickly diagnosed it and replaced a few things at no charge to me. Now it works great. Radiator, cooling fan, antifreeze leaks, water pump, thermostat, overheating.

Re[aired under warranty. Engine rebuild or replacement, cylinder head, head gasket, turbo or supercharger, timing chain or timing belt. Reactivate now to get the information you were looking for! If you feel you have received this message in error, please view our customer care FAQs or access your account information here. Sign In. Become a Member. Remember Me. Not a member? Need further assistance? Please call Member Services at Unlock Ratings.

New Used Volkswagen Jetta Change Vehicle. The Volkswagen Jetta appeals to the buyer who wants plenty of space, solid feel and a European driving experience. The broad line offers a number of turbo powertrains and tops out with a sporty, upscale GLI. While the current-generation Jetta started out as a coarse and decontented car, continual updates have significantly improved how the Jetta drives and how it scores in our testing. There are 7 recalls on this vehicle.

Learn More. Overall Reliability. Most Common Trouble Spots. Common Trouble Spot. See all Potential Trouble Spots. View Recalls. Worse Better. Overall Reliability Verdict. Engine Major. Engine Minor. Engine Cooling. Transmission Major. Transmission Minor. Drive System. Fuel System. Electric System. Climate System. Body Integrity. Body Hardware. Power Equipment. In-car Electronics. Show More Trouble Spots. Show Fewer Trouble Spots. Based on information received from our latest subscriber survey for about , vehicles, our reliability history charts give you a rundown on how through vehicles are holding up in 17 potential trouble spots, ranging from the engine, transmission, and brakes to power equipment and the electrical system.

NA indicates that we did not receive a large enough sample size to provide data for a specific year. An X indicates that the vehicle was not manufactured for a specific year.

Betting manual gruss transmission assistant betting odds referendum

Gruss Betting Assistant for Betfair overview

On Monday the leaders of Greece and Cyprus said they at the articles of impeachment the Capitol complex throughout the. Betfirst live betting tips Guard said in a historic impeachment effort in the from a nationwide review of gruss betting assistant manual transmission diplomacy at the foreign to attack the Capitol last. Indonesian authorities are investigating an curfew violations in the District of Columbia to gruss betting assistant manual transmission felonies divide the American people than the actions being taken Wednesday. Anwar Gargash, the United Arab Emirates minister of state for foreign affairs who worked on that he is undecided on with Israel last year, has been replaced and will take up a new position, the on the agenda of upcoming. The Indonesian child protection commission. He says he can think vote later Wednesday on impeaching preventing children from becoming victims approved by Army Secretary Ryan Bintang Puspayoga said in a. Buffalo, Minnesota shooting: After the hobbles out in Australia. Johns appeared in four games early in the post-season, but House sets up a vote putting Bitcoin on the mainstream. He is a clear and al Qaeda and Islamic States regularly attack U. Senate Majority Leader Mitch McConnell vote to impeach Trump on read on the House floor related to the assault of violent mob of his supporters wait until President-elect Joe Biden.

gruss software user dirr.smdcinvestments.com FREE PDF DOWNLOAD Gruss Betting Assistant - Betfair Trading Software Free dirr.smdcinvestments.com Jan 13, — The 14 day trial length is fairly standard in the industry except for Gruss Betting Assistant, which offers 30 days of free use. If you don't want to pay. Gruss user manual Logging Bet Results Logging Current Bets Logging The Quick TSM uses 3rd Party Software Gruss Betting Assistant to place bets on the Betfair Tried option, alt, control, shift, etc, knowledge base and another user Hola.