Mitochondrial Monoamine Oxidase

Abstract

Abstract Pargyline (N-benzyl-N-methyl-2-propynylamine), known to react stoichiometrically and irreversibly with the mitochondrial monoamine oxidase of bovine kidney, involving simultaneously the enzyme's flavin component, (Hellerman, L., and Erwin, V. G. (1968) J. Biol. Chem. 243, 5234–5243), has been shown to inactivate by forming a stable adduct with the flavin residue. Thus, the reaction of pargyline with an equivalent quantity of oxidase produced bleaching of the flavin at 455 nm and the appearance of a strongly absorbing species at 410 nm. Use of excess [7-14C]pargyline gave a stable 14C-protein product (1.1 residues of inhibitor bound per enzyme equivalent.) Proteolysis of the 14C-labeled protein and chromatographic fractionation of the peptides resulting revealed that most of the 14C label was associated with the fraction containing the altered coenzyme now absorbing maximally at 398 nm with a molar absorptivity of approximately 29,000 cm-1 m-1. Given here also is a procedure for the preparation of monoamine oxidase with twice the activity of the enzyme obtained previously (Erwin, V. G., and Hellerman, L. (1967) J. Biol. Chem. 242, 4230–4238). Based on estimates of enzyme concentration obtained by discontinuous titrations of the enzyme with pargyline, the respective values for the equivalent weight and the catalytic center activity (turnover) were calculated to be 109,000 and 525 for oxidation of benzylamine in air at pH 7.6 and 37°.