Biochemical characterization of the active site of brain monoamine oxidase.

Abstract

Until recently little was known concerning the chemical details of the mechanism of interaction of flavin-linked mitochondrial membrane bound monoamine oxidase (MAO) with its substrates and inhibitors. Substrates which have enzymes as their targets have been valuable in elucidating active site residues and structural features. Acetylenic amines as exemplified by clorgyline, deprenyl and pargyline are called 'suicide inhibitors' because an irreversible inhibitor is formed by the action of MAO from a relatively innocuous compound which acts as a substrate. These inhibitors can selectively inactivate MAO 'type A' and/or 'type B'. MAO isolated in homogeneous form from liver or kidney contains 1 mole of covalently bound coenzyme, cysteinyl-flavin, per mole enzyme. The flavin is bound to a pentapeptide via the thio-ether of cysteine at the 8 alpha-position of the isoalloxazine. A comparison of the inhibitory effects of clorgyline, deprenyl and pargyline on liver enzyme preparations from bovine or rat have confirmed our expectation that these irreversible inactivators form the same type of adduct with the cysteinyl-flavin active site of MAO 'type A' and 'type B', and that binding is stoichiometric at the N-5 of the covalently bound flavin in a flavocyanine linkage. Substrates protect from inhibition. In contrast to the reported observation of Tipton (39), pig brain mitochondrial MAO purified by two alternative methods contains cysteinyl-flavin in substantial amounts. The turnover number of enzyme from brain per mole of cysterinyl-flavin in apparently homogeneous samples is nearly the same as that of highly purified kidney and liver enzyme. Thus it is apparent that brain MAO also contains cysteinyl-flavin at the active center and therefore it is expected that acetylenic as well as hydrazine inhibitors form the same linkage with the flavin moiety as that formed with enzyme from peripheral tissues. A specific inhibitor for the deamination and potentiation of dopamine formed in the brain of Parkinsonian patients after treatment with L-Dopa has been regarded desirable. Deprenyl, a selective MAO 'type B inhibitor without the 'cheese effect', is the most potent inactivator of human brain MAO, and clinical results show that the drug is very useful in the treatment of Parkinson's disease and depression.