This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Eisenberg, R. C. Search for Related Content PubMed PubMed Citation Articles by Eisenberg, R. C.

Restoration of Deoxycholate-Disrupted Membrane Oxidases of Micrococcus lysodeikticus¹

^a Department of Biology, Western Michigan University, Kalamazoo, Michigan 49001

ABSTRACT

Membrane-associated L-malate and reduced nicotinamide adenine dinucleotide (NADH) oxidase complexes of Micrococcus lysodeikticus were inactivated with deoxycholate. Reactivation of NADH oxidase by addition of Mg²⁺ occurred in these detergent-membrane mixtures, but reactivation of L-malate oxidase did not occur in the presence of deoxycholate. Removal of detergent by gel filtration allowed Mg²⁺-dependent restoration of both L-malate and NADH oxidases. Maximal NADH and L-malate oxidase restoration required 10 min and 40 min, respectively, at 30 mM MgSO₄. Maximal restoration of both oxidases required at least 12 mM MgSO₄ in an incubation period of 1 hr. Reduced-minus-oxidized difference spectra of Mg²⁺-restored membrane oxidases showed participation of cytochromes b, c, and a when either L-malate or NADH served as reductant; addition of dithionite did not increase the - and ß-region absorbancy maxima of these hemoproteins when restored membranes were first reduced with the physiological substrates L-malate or NADH. Not all divalent cations tested were equally effective for reactivation of both oxidases. L-Malate oxidase was restored by both Mn²⁺ and Ca²⁺. NADH oxidase was not activated by Mn²⁺ and only slightly stimulated by Ca²⁺. Separation of deoxycholate-disrupted membranes (detergent removed) into soluble and particulate fractions showed that both fractions were required for Mg²⁺-dependent oxidase activities. Electron micrographs indicated conditions of detergent treatment did not destroy the vesicular nature of protoplast ghost membranes.

¹ Presented in part at the 71st Annual Meeting of the American Society for Microbiology, Minneapolis, Minn., 2–7 May 1971.