Flavoenzyme catalysis: substrate-competitive inhibition of D-amino acid oxidase.

In an earlier paper from this laboratory (1) there were described the characteristics of competitive inhibition resulting from the interference of certain aromatic compounds with the association of flavin adenine dinucleotide and the separated protein of n-amino acid oxidase. Included among the inhibitors were auramine, quinine, and various other quinolines and anilines. We now present our findings which relate to the structural requirements for some well defined substrate-competitive processes. The antimetabolic action of benzoate and cinnamate has been demonstrated by their ability to interfere with the oxidation of such metabolites as butyric, crotonic, and acetic acids (2-5) as well as certain Land Damino acids (6-8) in kidney and liver slices and homogenates. More recently we reported that benzoate and cinnamate can effect metabolic “shunts” in slices of various organs of the guinea pig and rat. In particular, cinnamate was shown to suppress the oxygen consumption of these tissues and to elicit a marked glycolytic production of lactate. Moreover, the inhibition of aerobic activity was found to be reversible and could be interpreted in terms of a competitive mechanism, embarrassing the oxidation of fatty acids and amino acids (9). With respect to the action of benzoate on an isolated enzyme system such as n-amino acid oxidase, Klein and Kamin (8) suggested that the inhibition was related to the formation of a benzoic acid-enzyme complex. In more detailed kinetic studies of this antagonism of amino acid oxidation by benzoate, we found that the action of this inhibitor is primarily substrate-competitive (1, 10). This was confirmed by Bartlett in his studies on various substituted benzoates (11). More recently, Klein has reported that certain heterocyclic acids with aromatic properties also can inhibit this system competitively (12). In the present paper, evidence will be presented that substrate-competitive inhibition of n-amino acid oxidase is not limited to compounds