Succinate-linked nicotinamide-adenine dinucleotide reduction coupled with the aerobic oxidation of reduced tetramethyl-p-phenylenediamine in submitochondrial particles

The energy-dependent reduction of NAD⁺ by succinate catalyzed by submitochondrial particles derived from beef heart is demonstrated to be supported energetically not only by the addition of ATP but also by a high-energy intermediate generated by the flow of electrons through the cytochrome oxidase portion of the respiratory chain. As substrate for the energy generation is used ascorbate and tetramethyl-p-phenylenediamine. Phenazinemethosulfate but not cytochrome c can replace the phenylenediamine. The reduction does not require the addition of Mg²⁺, phosphate or nucleotides, it is not inhibited by EDTA or oligomycin and by arsenate only in the absence of oligomycin, when the generated high-energy intermediate is the energy source. Rotenone is inhibitory whereas antimycin A is an obligatory addition to avoid reoxidation of the formed NADH.