Proteome analysis of Streptococcus mutans metabolic phenotype during acid tolerance

Abstract

Two-dimensional gel electrophoretic analysis of the proteome of Streptococcus mutans grown at a steady state in a glucose-limited anaerobic continuous culture revealed a number of proteins that were differentially expressed when the growth pH was lowered from pH 7?0 to pH 5?0. Changes in the expression of metabolic proteins were generally limited to three biochemical pathways: glycolysis, alternative acid production and branched-chain amino acid biosynthesis. The relative level of expression of protein spots representing all of the enzymes associated with the Embden-Meyerhof-Parnas pathway, and all but one of the enzymes involved in the major alternative acid fermentation pathways of S. mutans, was identified and measured. Proteome data, in conjunction with end-product and cell-yield analyses, were consistent with a phenotypic change that allowed S. mutans to proliferate at low pH by expending energy to extrude excess H + from the cell, while minimizing the detrimental effects that result from the uncoupling of carbon flux from catabolism and the consequent imbalance in NADH and pyruvate production. The changes in enzyme levels were consistent with a reduction in the formation of the strongest acid, formic acid, which was a consequence of the diversion of pyruvate to both lactate and branched-chain amino acid production when S. mutans was cultivated in an acidic environment.

Up-regulation of competence- but not stress-responsive proteins accompanies an altered metabolic phenotype in Streptococcus mutans biofilms

“…Notwithstanding, neither this nor previous 2-DGE analyses (Len et al, 2003(Len et al, , 2004b have identified the integral cytoplasmic membrane glucose transporters EII man and EII glc , or the glucose permease of S. mutans required for uptake of glucose (Cvitkovitch et al, 1995;Vadeboncoeur et al, 1991) due to their hydrophobic structure. As a consequence, any change that the level of expression of these transporters may have on glucose uptake and subsequent metabolic rate cannot be surmised.…”

“…Alterations in the fermentation phenotype of the S. mutans biofilm It is logical to hypothesize that any slowing of the growth rate in mature biofilms might be reflected in a down-turn in the level of metabolic enzymes, even though care should be taken in interpreting such a reduction as implying a corresponding down-turn in the flux of metabolites (Len et al, 2004b). Notwithstanding, neither this nor previous 2-DGE analyses (Len et al, 2003(Len et al, , 2004b have identified the integral cytoplasmic membrane glucose transporters EII man and EII glc , or the glucose permease of S. mutans required for uptake of glucose (Cvitkovitch et al, 1995;Vadeboncoeur et al, 1991) due to their hydrophobic structure.…”

Identification of genes associated with mutacin I production in Streptococcus mutans using random insertional mutagenesis

“…The dysfunction of AdhE may cause shunting of pyruvate towards the production of formate. Formic acid is a strong acid, which may contribute to a greater production of intracellular H + (Len et al, 2004). This possible internal acidification may create a severe stress signal leading to the abrogation of mutacin production.…”

“…CiaH is the histidine kinase of a two-component signal-transduction system, CiaH/R (Giammarinaro et al, 1999;Zähner et al, 2002), while LuxS is responsible for the synthesis of the interspecies cell signalling molecule autoinducer 2 (AI-2) (Schauder et al, 2001;Xavier & Bassler, 2003). In addition to regulating mutacin I production, both CiaH and LuxS were also found to be involved in regulating other cellular functions in S. mutans such as biofilm formation and stress tolerance (Len et al, 2004;Merritt et al, 2003;Qi et al, 2004). As a preliminary attempt to understand the control of mutacin I The pZero-2 cloning vector (Invitrogen) was used to facilitate library construction in E. coli, and as a suicide vector in S. mutans.…”

Ion-paired RP-HPLC for determining fermentation end products of Streptococcus mutans grown under different conditions

“…By the appropriate use of the acetate and ethanol branches of the PFL pathway, S. mutans can readily adjust NADH oxidation to the actual need. Without further anabolic diversion of the fi nal products, the amount of formic acid produced by glycolysis is equivalent to the total amount of acetic acid and ethanol 6 . An important characteristic which distinguishes S. mutans from other oral streptococci is its ability to ferment sugar alcohols such as sorbitol, and mannitol.…”