Amino Acid Oxidase, L-

I.U.B.: 1.4.3.2
L-amino-acid: O₂ oxidoreductase (deaminating)

L-amino acid oxidase (LAO) catalyzes the oxidative deamination of a number of L-amino acids:

LAO from snake venom has received the most attention: it has been reviewed by Meister and Wellner (1963) and Bright and Porter (1975), the latter dwelling especially on kinetics involved.

The enzyme is important for the purification and determination of certain amino acids and the preparation of a-keto acid. Nicholson and Kim (1975), Auricchio et al. (1971), and Donlon and Fottrell (1971) report on its use in assaying peptidase activity -- the rate of H₂O₂ is determined colorimetrically by its oxidation of o-dianisidine in the presence of peroxidase. Holme and Goldberg (1975) describe a spectrophotometric assay system for LAO which may be of further interest: NH₃ released is coupled with the NADH-dependent reductive amination of 2-oxoglutarate catalyzed by glutamate dehydrogenase.

Guilbault and Hrabankova (1970) describe an enzyme electrode using LAO for measuring L-amino acids. See also Tran-Minh and Brown (1975).

Characteristics of LAO from Crotalus venom:

Molecular weight: 140,000 (DeKok and Rawitch 1969).

Composition: A glycoprotein consisting of two differing subunits of approximately MW 70,000 combined in unequal amounts. Three electrophoretically different isozymes result from different combinations of the two kinds of subunits. There are two FAD molecules per mole of holoenzyme.

Optimum pH: Approximately 7.5 for leucine (Wellner and Meister, 1960).

Extinction coefficient: = 17.9.

Specificity: Absolute for L-isomers. Substrates are L-isomers of leucine, isoleucine, norleucine, α-amino butyric acid, phenylalanine, tyrosine, tryptophan norvaline, methionine, histidine and citrulline. Histidine and tyrosine cannot be determined in an LD-mixture (Malmstadt and Hadjiioannov 1963). Methylene blue may be used as an electron acceptor. L-serine, threonine, aspartic acid, glutaric acid, lysine and ornithine are only slightly attacked.

Inhibitors: Aromatic carboxylates inhibit the enzyme competitively (DeKok and Veeger, 1968). See also Sankar et al . (1975).

Stability: The enzyme is stable in solution for months when refrigerated. Substrate and absence of oxygen protect at elevated temperatures. The enzyme may be reversibly inactivated by incubation at 38°C in phosphate buffer, pH 7.5 (Wellner, 1966). Curti et al. (1968) report reversible inactivation upon freezing.