title: Fructose-1,6-diphosphate-dependent lactate dehydrogenase from a cariogenic streptococcus: purification and regulatory properties.
abstract: The lactate dehydrogenase (LDH) from Streptococcus mutans NCTC 10449 is under stringent metabolic control. The partially purified enzyme was specifically activated by high concentrations of fructose-1,6-diphosphate (FDP) and was inhibited by adenosine triphosphate. There appeared to be at least two binding sites for the activator which interacted in a cooperative manner. The interaction between the FDP sites was independent of the pH of the assay system, although the relative affinity of the enzyme for the activator was influenced by pH. There also appeared to be at least two pyruvate binding sites on the S. mutans LDH with some cooperative interaction between them, and the interaction between these sites was also independent of the hydrogen ion concentration. Two pyruvate analogues had different effects on the interaction of pyruvate with the LDH. One of the analogues, alpha-ketobutyrate, stimulated enzyme activity at limiting pyruvate concentrations, but had no significant effect at saturating concentrations of the substrate. The net effect of alpha-ketobutyrate was to shift the pyruvate saturation curve from sigmoidal to hyperbolic and to decrease the Hill coefficient from about 2.0 to 1.0. The other pyruvate analogue, oxamate, inhibited enzyme activity at all pyruvate concentrations but had no effect on the sigmoidal nature of the pyruvate saturation curve or on the apparent kinetic order of the reaction with respect to substrate. These results suggested that there may be two types of pyruvate binding sites on the LDH from S. mutans. Other kinetic properties of the S. mutans NCTC 10449 enzyme were studied and compared with those exhibited by the LDH from several other strains of the organism.
mesh_terms: D000255:Adenosine Triphosphate / Q000494:pharmacology; D000645:Ammonium Sulfate; D001665:Binding Sites; D002474:Cell-Free System; D011232:Chemical Precipitation; D002848:Chromatography, DEAE-Cellulose; D002852:Chromatography, Ion Exchange; D003731:Dental Caries / Q000382:microbiology; D004789:Enzyme Activation; D004794:Enzyme Repression; D005632:Fructose / Q000494:pharmacology*; D005636:Fructosephosphates / Q000378:metabolism / Q000494:pharmacology; D006600:Hexosephosphates / Q000494:pharmacology*; D006358:Hot Temperature; D006851:Hydrochloric Acid; D006863:Hydrogen-Ion Concentration; D007770:L-Lactate Dehydrogenase / Q000302:isolation & purification / Q000378:metabolism*; D009243:NAD / Q000378:metabolism; D011773:Pyruvates / Q000378:metabolism / Q000494:pharmacology; D013291:Streptococcus / Q000201:enzymology*
chemical_list: D005636:Fructosephosphates; D006600:Hexosephosphates; D011773:Pyruvates; D009243:NAD; D005632:Fructose; D000255:Adenosine Triphosphate; D007770:L-Lactate Dehydrogenase; D006851:Hydrochloric Acid; D000645:Ammonium Sulfate