Studies on metmyoglobin reducing enzyme systems in the muscle of blue white dolphin. I. Purification and properties of diaphorase.

Publisher Website
Google Scholar
Abstract
Diaphorase was isolated from the skeletal muscle of blue white dolphin by DEAE-cel-lulose column chromatography, Sephadex G-100 gel filtration, starch block electrophoresis, etc. The diaphorase obtained showed an apparent homogeneity in both ultracentrifugal and electrophoretical analyses. Its sedimentation and diffusion coefficients were 6.28 S and 4.95× 10-7cm2 sec-1, respectively, and a molecular weight (MS, D) of 120, 000 was calculated. This enzyme was found to contain two moles of FAD per mole. NADH2, but not NADPH2, served as an electron donor. Reducing activities of dolphin diaphorase for metmyoglobin (MMb), dichlorophenol indophenol (DPIP), lipoamide, and lipoic acid were 0.55, 0.81, 11.5, and 1.20 U per mg protein, respectively, The Michaelis constants in these reactions were calculated to be 2.2×10-5M, 1.3×10-5M, 1.0×10-3M, and 4.2×10-3M in the above order, respectively. The lipoamide reducing activity was strongly inhibited by dithiol inhibitors. On the contrary, the MMb and DPIP reducing activities were activated by these inhibitors. Both activities were, however, effectively inhibited by PCMB. It was demonstrated that the MMb reducing activity is accelerated by lipoamide and lipoic acid to the same extent as by methylene blue.

This publication has 0 references indexed in Scilit: