OXIDATION OF REDUCED NICOTINAMIDE-ADENINE-DINUCLEOTIDE IN RHODOSPIRILLUM-RUBRUM .3. PROPERTIES OF A NADH DEHYDROGENASE SOLUBILIZED FROM ELECTRON TRANSPORT PARTICLES

The properties of a NADH dehydrogenase which was solubilized from the electron transport particulate fraction of R. rubrum by treatment with DOC were investigated. 1. The solubilized activity was further purified by DEAE-chromatography (total purification appr. 30fold). The purified enzyme preparation, because of its very low protein concentration, is extremely unstable. 2. The molecular weight of the solubilized enzyme, as determined by gelfiltration through Sephadex, is 75,000. 3. The solubilization increases the affinity of the enzyme for the electron aceptors, while the affinity for its substrate remains unchanged. 4. The enzyme is increasingly inactivated by dilution in buffer. 5. Incubation with the substrate NADH rapidly inactivates the enzyme. This inactivation is specific (NADPH inactivates only at much higher concentrations, NAD and NADP are without effect), it is complete and also irreversible. No protection can be achieved. 6. 2×10-4M PCMB or mersalyl completely inhibit the enzyme, EMI is without effect. There is no evidence for an inhibition, by PCMB or mersalyl when the system is pretreated with NADH (SH-group of Tyler et al., 1965). 7. o-phenanthroline is the only chelating agent found to interact with the NADH dehydrogenase. With only quantitative differences the properties given under 4.-7. are also shown by the membrane bound activity of NADH dehydrogenase, thus indicating a mild attack of DOC on the membranal structures and an easy, release of the enzyme. © 1969 Springer-Verlag.