Mitochondrial malate dehydrogenase of bovine cerebrum. Characterization and mechanisms of inhibition by silver ions.
Attempts were made to characterize mitochondrial malate dehydrogenase [L-malate: NAD+ oxidoreductase, EC 1.1.1.37] (M-MDH) purified from bovine cerebrum and to elucidate the mechanisms responsible for inhibition of the enzymic activity by Ag+. The molecular weights of the native enzyme and its subunits were 54,000-55,000 and 30,000-32,000, respectively. In general, the physiochemical and catalytic properties of bovine cerebral M-MDH was not very different from those of other corresponding mammalian enzymes. Incubation of the enzyme with Ag+ caused the loss of equivalent amounts of sulfhydryls with a parallel decrease of the enzymic activity. When the enzyme was exposed to 2-, 3.5-, and 5-fold molar excesses of Ag+, the enzymic activity showed an initial rapid fall and a subsequent slow restoration to a partially inactivated level (60-70, 45-50, and 15-20% of an untreated control, respectively), while the alpha-helical content of the enzyme fell exponentially with time. A 7-fold molar excess of Ag+ reduced both the enzymic activity and the alpha-helical content to a much greater degree and no restoration of the enzymic activity was observed. The Km values of Ag+-inactivated enzyme for NADH and oxaloacetate were the same as those of the native enzyme. The data suggest that Ag+ could inhibit enzymic activity both by reducing the structural regularity of the enzyme molecule and by attacking sulfhydryl groups necessary for the catalytic activity of bovine cerebral M-MDH.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001921:Brain / Q000201:enzymology*; D002417:Cattle; D004305:Dose-Response Relationship, Drug; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D046911:Macromolecular Substances; D008291:Malate Dehydrogenase / Q000037:antagonists & inhibitors*; D008928:Mitochondria / Q000201:enzymology*; D008970:Molecular Weight; D009243:NAD / Q000494:pharmacology; D010071:Oxaloacetates / Q000494:pharmacology; D012834:Silver / Q000494:pharmacology*; D013439:Sulfhydryl Reagents / Q000494:pharmacology
D000596:Amino Acids; D046911:Macromolecular Substances; D010071:Oxaloacetates; D013439:Sulfhydryl Reagents; D009243:NAD; D012834:Silver; D008291:Malate Dehydrogenase