Oxidative deamination of sulfur amino acids by bacterial and snake venom l-amino acid oxidase

Abstract

We have investigated the oxidative deamination for all available sulfur amino acids by the crystalline snake venom l -amino acid oxidase from Crotalus terrificus terrificus and a particulate bound oxidase from an atypical Proteus rettgeri. Specific activity, pH optimum, Km and Vmax values were determined for each sulfur amino acid. For the bacterial enzyme the affinity for the various substrates at pH 7.5 in decreasing order is l -homocysteine, l -cysteine, l -methionine, l -homocystine, S-ribosyl- l -homocysteine, S-adenosyl- l -homocysteine, S-adenosyl- l -methionine, methionine sulfoxide, l -djenkolic acid, S-aderosyl- l -homocysteine sulfoxide, and l -cystine. No measurable activity was observed with cysteic acid, homocysteic acid, lanthionine, cystathionine, or S-ribosyl- l -homocysteine sulfoxide. The snake venom enzyme had a somewhat more limited specificity. There was no measurable activity with any of the sulfoxides, S-adenosyl- l -methionine, cysteic acid, homocysteic acid, lanthionine, or cystathionine. The affinity for the various substrates at pH 7.5 in decreasing order is l -methionine, l -homocystine, l -homocysteine, S-adenosyl- l -homocysteine, S-ribosyl- l -homocysteine, l -djenkolic acid, l -cysteine, and l -cystine. The keto acids produced from the various substrates with both enzymes were identified by chemical analysis and by thin-layer chromatography of the free keto acid or their dinitrophenylhydrazones against known standards. All the resultant keto acids appear to correspond to their parent compounds. Homocystine was found to be completely oxidized to 4,4′-dithio-bis (2-ketobutyric acid). S-Adenosyl- l -methionine, which is resistant to attack by most enzymes including snake venom l -amino acid oxidase, was readily oxidized to S-adenosyl-α-keto-γ-methiolbutyrate by the bacterial enzyme. The mercapto-α-keto acids derived from homocysteine and cysteine were found to be unstable resulting in the liberation of hydrogen sulfide and α-ketobutyrate or pyruvate, respectively.