Acid production by streptococci growing at low pH in a chemostat under anaerobic conditions

Abstract

Streptococcus mutans and other oral streptococci were grown in continuous culture under strictly anaerobic conditions. When the cultural pH was kept at 7.0, the main acid products were formate and acetate, as reported previously. However, more lactate was produced at pH values of 5.5 or 6.0, with a concomitant decrease in formate and acetate production. This change in fermentation products could partly be ascribed to a change in intracellular pH and difference in the pH optima between pyruvate formate-lyase (PFL) and lactate dehydrogenase (LDH). At extracellular pH values of 7.0 and 5.5, the intracellular pH values of S. mutans NCIB 11723 were 7.5 and 6.6, respectively. The pH optima of PFL and LDH were 7.8 and 5.5-6.3, respectively. The cells had also a larger amount of LDH during growth at pH 5.5 than at pH 7.0.

Proteome analysis of Streptococcus mutans metabolic phenotype during acid tolerance

“…Studies of changes in the levels of protein expression, along with measurement of the final by-products of carbon utilization, Y Glc and Y ATP , have confirmed previous findings that streptococci tolerate growth at pH 5?0 by increasing their ability to extrude H + , while modulating the production of acid fermentation by-products to reduce the level of H + production (Hamilton, 1984;Iwami et al, 1992;Miyagi et al, 1994). The results further suggest that S. mutans utilizes branched-chain amino acid biosynthesis to lower the amount of reducing equivalents in the form of NAD(P)H that would otherwise be converted to acidic by-products, and by so doing helps buffer the cytoplasm by producing NH 3 .…”

“…The levels of the final three enzymes in glycolysis at pH 5?0 were consistent with a reduced pool of phosphoenolpyruvate, since the reduced levels of enolase, in conjunction with an overall 2?8-fold increase in the four isoforms of pyruvate kinase, would be expected to convert 2-phosphoglycerate to pyruvate without any build-up of phosphoenolpyruvate. This idea is supported by the measurement of the four intermediates, 3-phosphoglycerate, 2-phosphoglycerate, phosphoenolpyruvate and pyruvate, in S. mutans grown at pH 5?5 and D=0?15 h 21 , where the levels of 3-phosphoglycerate and phosphoenolpyruvate decrease, that of 2-phosphoglycerate remains static and the amount of pyruvate increases (Iwami et al, 1992). This is further supported by another study, where a fall in intracellular pH was shown to significantly increase the pyruvate : phosphoenolpyruvate ratio (Iwami & Yamada, 1980).…”

“…These values are similar to reports of a total 1?7-fold increase observed on 2-DGE gels for the three isoforms of Llactate dehydrogenase in S. mutans and the five isoforms in S. oralis when grown without pH control in batch culture (Wilkins et al, 2001(Wilkins et al, , 2002. The increase in L-lactate dehydrogenase isoforms observed in our study is unlikely to account for the 127-fold increase in the amount of L-lactic acid produced under acidic conditions (Table 2), despite the fact that L-lactate dehydrogenase has a broad optimum for catalytic activity in the acidic pH range 5?0-6?2, similar to the pH range 6?0-6?5 of the cytoplasm of cells growing at pH 5?0 (Dashper & Reynolds, 1990;Hamilton, 1990;Iwami et al, 1992). The more likely explanation is that the enzyme is regulated by glycolytic intermediates at the lower growth pH (Yamada & Carlsson, 1975b;Takahashi et al, 1982;Yamada, 1987).…”

“…Previous analysis of metabolic intermediates formed in the first half of the Embden-Meyerhof-Parnas pathway indicates that the level of glucose 6-phosphate is slightly elevated when S. mutans is grown in continuous culture at pH 5?5 and D=0?15 h 21 , while the levels of all other intermediates are essentially the same, up to and including dihydroxyacetone phosphate and glyceraldehyde 3-phosphate (Iwami et al, 1992). On this basis, the doubling of glucose-6-phosphate isomerase (Gpi) and an overall 1?5-fold increase in the level of the three isoforms of fructose-1,6-bisphosphate aldolase (Fba), similar to that previously observed in batch culture (Wilkins et al, 2002; Fig.…”

“…However, as glucokinase has a pH optimum of approximately 8?0 (Porter et al, 1982), its relative activity and stability are affected by acidification of the cytoplasm at a growth pH of 5?0, where the internal pH would be less than 6?5 (Dashper & Reynolds, 1990;Hamilton, 1990;Iwami et al, 1992). Since the optimal pH for glycolysis appears to be 7?0 (Dashper & Reynolds, 1992), the reduction in cytoplasmic pH most likely explains why glucokinase is a dPart of the complement of random landmark proteins used to align gels and to calculate observed pI and M r, in accordance with the requirements of the z3 software.…”