Increase in tryptamine oxidation activity of hog kidney mitochondrial monoamine oxidase by treatment with triton-X 100 and sodium cholate.

Monoamine oxidase (MAO) [EC 1.4.3.4 monoamine: oxygen oxidoreductase (deami nating)] has been shown to be firmly bound to the mitochondrial membrane (1-4), and its solubilization causes changes in its properties (5-7). This paper describes the solubilization of MAO from mitochondria of hog kidney cortex by a mixture of Triton-X 100 and sodium cholate and changes in the substrate specificities of this enzyme resulting from its solubilization. Hog kidney cortex was homogenized with 4 volumes of 0.25 M sucrose containing 0.1 M potassium phosphate buffer (pH 7.4). The homogenate was centrifuged at 600 x g for 10 min, and the resulting supernatant was recentrifuged at 8,500 x g for 30 min. The resulting precipitate was suspended in 9 volumes of sucrose buffer and used as mito chondrial MAO (protein 6.6 mg/ml). Another portion of the precipitate was suspended in 9 volumes of 0.1 M potassium phosphate buffer, pH 8.4, containing 1 % Triton-X 100 and 0.5% sodium cholate by a slight modification of the method of Minamiura and Yasunobu (8). This suspension was stirred for 4 hr at 4'C and then kept in a deep freezer for 24 hr. The frozen enzyme was then thawed at room temperature and used as Triton-X and cholate treated MAO. MAO activity was determined manometrically and expressed as oxygen uptake (ill 02) in 60 min at 38'C under a gas phase of 100% oxygen (9). Protein contents were determined by the modified biuret method using bovine albumin as a standard (10). The mitochondrial MAO of hog kidney showed the highest activity with tyramine (119 /eI 02/hr/0.7 ml enzyme) followed in order of decreasing activity by tryptamine (96 ,fl 02), isoamylamine (87 /el 02), serotonin (76 Itl 02), octopamine (43 ul 02), benzylamine (42 rcl 02) and beta-phenyl ethylamine (20 itl 02), as shown in Fig. 1 (left). However, Triton-X and cholate treated MAO showed highest activity with tryptamine (119 /11 02/hr/0.7 ml enzyme) followed by tyramine (66 ul 02), isoamylamine (60 /dl 02), benzylamine (40 ,al 02), serotonin (39 /el 02), octopamine (23 ,ul 02) and beta-phenyl ethylamine (4 tI 02), as shown in Fig. 1 (right). MAO activity in hog kidney with tryptamine as substrate was increased about 30% by treatment of the mitocondria with Triton-X 100 and sodium cholate, while its activities with the other substrates were all decreased by this treatment. In addition, it was found that the increase in tryptamine oxidation …