Evidence for the importance of cysteine and arginine residues in Pseudomonas fluorescens UK-1 pantoate dehydrogenase.
Homogeneous pantoate dehydrogenase (D-pantoate:NAD+ 4-oxidoreductase, EC 1.1.1.106) was shown to be sensitive to inactivation by p-chloromercuribenzoate (100 microM), 5,5'-dithiobis(2-nitrobenzoic acid) (1 mM), iodoacetic acid (1 mM) and phenylglyoxal (5.3 mM). Potassium D-pantoate and NAD protected against inactivation by p-chloromercuribenzoate, 5,5'-dithiobis (2-nitrobenzoic acid) and iodoacetic acid. NAD and D-pantoate also provided substantial protection against inactivatrion by phenylglyoxal. Titration of the sulphydryl groups by 5,5'-dithiobis(2-nitrobenzoic acid) and incorporation of [14C]carboxymethyl revealed that there are two cysteine residues which are modified and one of those is essential for activity. In the presence of NAD and D-pantoate, incorporation of [14C]phenylglyoxal was decreased by 0.42 mol/mol of subunit.
D000429:Alcohol Oxidoreductases / Q000037:antagonists & inhibitors*; D001120:Arginine / Q000378:metabolism*; D002729:Chloromercuribenzoates / Q000494:pharmacology; D003545:Cysteine / Q000378:metabolism*; D004228:Dithionitrobenzoic Acid / Q000494:pharmacology; D006885:Hydroxybutyrates / Q000037:antagonists & inhibitors / Q000494:pharmacology; D007461:Iodoacetates / Q000494:pharmacology; D009243:NAD / Q000494:pharmacology; D010658:Phenylglyoxal / Q000494:pharmacology; D011551:Pseudomonas fluorescens / Q000201:enzymology*
D002729:Chloromercuribenzoates; D006885:Hydroxybutyrates; D007461:Iodoacetates; C016031:pantoic acid; D009243:NAD; D001120:Arginine; D004228:Dithionitrobenzoic Acid; D000429:Alcohol Oxidoreductases; C022316:pantoate dehydrogenase; D003545:Cysteine; D010658:Phenylglyoxal