Energy coupling mechanisms in host-grown Mycobacterium lepraemurium.

Energy coupling mechanisms of Mycobacterium lepraemurium isolated from Sprague-Dawley rats lepromata were investigated. Cell-free extracts catalyzed phosphorylation coupled to the oxidation of generated NADH, added NADH, and succinate yielding P/O ratios of approximately 0.8, 0.6, and 0.4, respectively. Ascorbate oxidation alone or in the presence of cytochrome c or N,N,N',N'-tetramethyl-p-phenylenediamine was not coupled to ATP synthesis. The oxidative phosphorylation was completely uncoupled by 2,4-dinitrophenol, 2,6-dibromophenol, pentachlorophenol, m-chlorocarbonylcyanide phenylhydrazone, dicumarol, and gramicidin at concentrations which did not cause any inhibition of oxygen uptake. While the NADH oxidation and associated phosphate esterification was markedly sensitive to rotenone and other flavoprotein inhibitors, these inhibitors had no effect, however, on the phosphorylation coupled to succinate oxidation. The respiratory chain inhibitors such as antimycin A or 2-n-heptyl-4-hydroxyquinoline-N-oxide, and cyanide were the potent inhibitors of the phosphorylation associated with the oxidation of NADH and succinate. The ATP formation coupled to the oxidation of NADH and succinate was also inhibited by oligomycin as well as by the thiol-binding agents, p-hydroxymercuribenzoate and N-ethylmaleimide. The results indicated that NADH and succinate oxidation by in vivo grown M. lepraemurium was mediated by oxidative enzymes involving first and second energy coupling sites.