Studies of monoamine oxidases: I. Purification and properties of the rabbit liver mitochondrial enzyme

Abstract

1. 1. Rabbit liver mitochondrial monoamine oxidase (monoamine:O 2 oxidoreductase (deaminating), EC 1.4.3.4) was purified approx. 270 fold. The purification procedure included isolation of the mitochondrial fraction, solubilization by sonication in the presence of 0.4% isooctylphenoxypolyethyoxyethanol, and centrifugation at 12 000 × g max for 30 min. The supernatant was then fractionated successively on DEALE-cellulose, Biogel P-300, DEAE-Sephadex A-50, and hydroxylapatite columns. 2. 2. Kynuramine was used both to measure enzyme activity in following the course of purification and to carry out kinetic studies. The K m value of kynuramine was 77 μM at pH 7.4. The pH optimum of the enzyme was about 8.4 with kynuramine and about 9.2 with another substrate, m-iodobenzylamine. 3. 3. Highly purified material was used to study some of its properties and the effect of chelating agents. The metal-chelating agents ( o-phenanthroline and neocuproine) displayed a dual effect on the activity of purified monoamine oxidase after preincubation of the enzyme with chelating agent at 37°. At low concentrations of chelator (2–100 μM), the enzymic activity was greater than without chelators, whereas at higher concentrations of these agents (400 μM and greater), enzyme inhibition took place.