Amino acid analogs, reaction with oxidases

Until recently, the amino acid oxidase reaction has been studied only in the direction of ammonia and a-keto acid formation. In the presence of air the reaction proceeds to completion and is essentially irreversible because of the reoxidation of the reduced flavoprotein by molecular oxygen [reaction (5)]. Meister and his associates 11, 12) have provided a clear demonstration of the reversibility of the amino acid oxidase reaction with D-amino acid oxidase (from sheep kidney) and L-amino acid oxidase (from snake venom). When an amino acid, ammonia, and the a-keto acid analog of a second amino acid are incubated with either amino acid oxidase under anaerobic conditions, the formation of the second amino acid is observed ... 

To understand the carbanion mechanism in flavocytochrome 62 it is useful to first consider work carried out on related flavoenzymes. An investigation into o-amino acid oxidase by Walsh et al. 107), revealed that pyruvate was produced as a by-product of the oxidation of )8-chloroalanine to chloropyruvate. This observation was interpreted as evidence for a mechanism in which the initial step was C -H abstraction to form a carbanion intermediate. This intermediate would then be oxidized to form chloropyruvate or would undergo halogen elimination to form an enamine with subsequent ketonization to yield pyruvate. The analogous reaction of lactate oxidase with jS-chlorolactate gave similar results 108) and it was proposed that these flavoenzymes worked by a common mechanism. Further evidence consistent with these proposals was obtained by inactivation studies of flavin oxidases with acetylenic substrates, wherein the carbanion intermediate can lead to an allenic carbanion, which can then form a stable covalent adduct with the flavin group 109). Finally, it was noted that preformed nitroalkane carbanions, such as ethane nitronate, acted as substrates of D-amino acid oxidase 110). Thus three lines of experimental evidence were consistent with a carbanion mechanism in flavoenzymes such as D-amino acid oxidase. 

---