ABSTRACT
The overall reaction mechanism of lactate oxidase was elucidated with the enzyme from Mycobacterium smegmatis, employing a combination of steady-state and stopped-flow kinetics. Lactate oxidase is also inactivated by reaction with fluorodinitrobenzene, and the presence of competitive inhibitors such as nitrate or ot-hydroxymalonate provided strong protection. Lactate oxidase from Mycobacterium phlei was found to be inactivated irreversibly in lutidine buffer by reaction with 2,3-butanedione or phenylglyoxal. The amino acid sequence of lactate oxidase is now available via the cDNA sequence of the cloned gene. The catalytic cycle of lactate oxidase is initiated by substrate dehydrogenation in the complex with oxidized enzyme. Lactate oxidase recognizes glycollate as a substrate and it catalyzes its oxidation in a fashion which is superficially similar, but in important details significantly different from that observed with lactate. Glycollate undergoes a dehydrogenation and an oxidative decarboxylation in the presence of oxygen, the main produets being formate, C02, and water.
