Studies of the energy-transfer system of submitochondrial particles. I. Competition between oxidative phosphorylation and the energy-linked nicotinamide-adenine dinucleotide transhydrogenase reaction
Lee, C.; Ernster, L.
European Journal of Biochemistry 3(4): 385-390
ISSN/ISBN: 0014-2956 PMID: 4296029 DOI: 10.1111/j.1432-1033.1967.tb19541.x
The relationship between oxidative phosphorylation and the energy-linked nicotinamide-adenine dinucleotide transhydrogenase reaction catalyzed by phosphorylating submitochondrial particles from beef heart has been investigated. The efficiency of the particles in exhibiting oxidative phosphorylation (P/O ratio) and NADH-linked NADP+ reduction (NADP+/O ratio) was measured, using succinate as oxidizable substrate and rotenone as inhibitor of Nadh oxidase, under conditions where the phosphorylating and transhydrogenating systems were operating one by one or in parallel. When succinate oxidation was limited by increasing concentrations of malonate, the P/O ratio remained unchanged in the absence of an active phosphorylating system, and decreased in its presence. The NADP+/O ratio increased with increasing concentrations of malonate, both in the absence and presence of an active phosphorylating system. The (P + NADP+)/O ratio in the combined system approached 2, i. e., the generally accepted maximal ∼/O ratio for succinate oxidase. Active transhydrogenation caused an increase in the apparent Km for Pi in the phosphorylating system from 1.7 to 4.5 m M, and active phosphorylation caused an increase in the apparent Km for Nadh in the transhydrogenating system from 21 to 40 μM. It is concluded that oxidative phosphorylation and the energy-linked transhydrogenase reaction are competing for a common nonphosphorylated high-energy intermediate generated by the respiratory chain. The kinetic evidence presented further suggests that the two systems interact with two different moieties of this high-energy intermediate.