Susan S. Chen

Date of Award


Document Type


Degree Name

Master of Science (MS)


Biomedical Sciences

First Advisor

John A. Duerre


The oxidative deamination of sulfur amino acids by a particulate L-amino acid oxidase obtained from an atypical Proteus rettgeri has been investigated. In addition to several sulfur amino acids tested here the enzyme catalyzes the oxidation of L-leucine, L-phenylalanine, L-tryptophan, L-arginine, L-tyrosine, L-histidine, L-ornithine, L-citrulline, L-lysine and L-isoleucine with a pH optimum at 7.5. Specific activity, pH optimum, Km and Vmax values were determined for each sulfur amino acid. The ordered rate of reaction at pH 7.5 for these amino acids were L-homocysteine, L-methionine, S-adenosyl-L-homocysteine, S-adenosyl-L-methionine, L-djenkolic acid, S-ribosyl-L-homocysteine, S-adenosyl-L-homocysteine sulfoxide, L-homocysteine, methionine sulfoxide, L-cysteine and L-cystine.

The particulate fraction contained sufficient catalase to catalyze the complete reaction as follows:

R-CH(NH2)-COOH + ½ O2 → R-CO-COOH + NH3

The products obtained with all these substrates, except for homocysteine and cysteine, were ammonia and keto acid. With the free sulfhydryl compounds oxygen consumption was disproportionately high in addition to ammonia and keto acids, hydrogen sulfide was liberated. No activity was observed under anaerobic conditions, nor did pyridoxal phosphate affect the rate of the reaction; therefore it appeared that hydrogen sulfide is a product of the reaction and not due to the presence of sulfhydrase. The crystalline snake venom L-amino acid oxidase from Crotalus terrificus terrificus was studied in comparison and found to catalyze a similar reaction.

The pH optimum for each substrate was determined either by measuring the rate of oxygen consumption or keto acid formation. The pH optimum for all sulfur amino acids with both enzymes appeared in the alkaline range. The shape of the pH curve was dependent on the individual substrate and enzyme used.

The keto acids produced from the various substrates with both enzymes were identified by thin-layer chromatography of the free keto acid or their dinitrophenylhydrazones. Except for homocysteine or cysteine all the resultant keto acids were found to correspond to the parent compound. Homocysteine was found to be completely oxidized to 4,4’-dithio-bis(2-ketobutyric acid) by chemical and elemental analysis. S-adenosyl-L-methionine which is resistant to attack by most enzymes including snake venom and rat kidney L-amino acid oxidase was readily oxidized to S-adenosyl-α-keto-γ-methiobutyrate by the bacterial enzyme.

The keto acids derived from homocysteine and cysteine by both enzymes were α-ketobutyrate and pyruvate respectively.