Nitromethane: A NOVEL SUBSTRATE FOR d-AMINO ACID OXIDASE

Abstract

Abstract d-Amino acid oxidase (EC 1.4.3.3) catalyzes the oxidation of nitromethane by O2 in the presence or absence of light to give formaldehyde, nitrite, and H2O2 as final products. The anaerobic reduction of the enzyme by nitromethane yields a species (E'r) the spectrum of which resembles, but is not identical with, that of free reduced enzyme. E'r reacts rapidly with O2 to form fully oxidized enzyme. The enzyme remains in the oxidized state during turnover at pH 8.3 because the rate of oxidation of E'r by O2 is 6 x 104 times faster than the rate of formation of E'r. The following scheme accommodates the observed kinetics of turnover and of the half-reactions. Eo (k1[CH3NO2])/→ E'r (k2[O2])/→ Eo + H2O2 Whether nitrite and formaldehyde (or a precursor such as HO—CH2—NO2) are released in the first or second step has not been established. In addition to undergoing oxidative turnover, nitromethane slowly produces an irreversibly inactivated species of the enzyme with none of the spectral characteristics of oxidized enzyme. Nitromethane interacts with free FAD, but only in the presence of light, to give an oxidizable species having a spectrum resembling that of FADH2.