Reconstitution of Micrococcus lysodeikticusReduced Nicotinamide Adenine Dinucleotide and l-Malate Dehydrogenases with Dehydrogenase-Depleted Membrane Residues: a Basis for Restoration of Oxidase Actitivies

Deoxycholate disruption of Micrococcus lysodeikticusprotoplast membranes resulted in solubilization of both l-malate and reduced nicotinamide adenine dinucleotide (NADH) dehydrogenase enzymes (substrate: 2,6-dichlorophenolindophenol oxidoreductases). Insoluble residues contained cytochromes of the b, c, and atype. Solubilized dehydrogenases were reconstituted with insoluble residues by treatment of disrupted membranes with magnesium ions. Most of the solubilized l-malate and NADH dehydrogenase activities were precipitated by magnesium ions independent of enzyme reconstitution with insoluble residues. Reconstituted dehydrogenases explained the mechanism for restoration of disrupted l-malate and NADH oxidase activities (4). Black light irradiation inhibited oxidase activities of both native and reconstituted membranes. These irradiated membrane oxidases were partially restored by exogenous napthoquinones [K2(20)and K2(50)] but not by CoQ(6). Reconstitution experiments showed that native membrane napthoquinone was retained in the insoluble residues of deoxycholate-disrupted membranes.