PROTON TRANSLOCATION LINKED TO THE ACTIVITY OF THE BI-TRANS-MEMBRANE ELECTRON-TRANSPORT CHAIN

Recently we have proposed and presented evidence suggesting the existe nce of a ''bi-trans-membrane'' electron transport chain, located at th e contact sites between outer and inner mitochondrial membranes, which can be utilized to promote either the oxidation of exogenous NADH in the presence of catalytic amounts of added cytochrome c or the reducti on of exogenous cytochrome c supported by the oxidation of respiratory substrates present inside the mitochondria. Here we show that the oxi dation of exogenous NADH is accompanied by a net alkalinization of the incubation medium preceded by a transient acidification phase. In oxy gen-pulse experiments, the alcohol oxidation (induced by the addition of alcohol dehydrogenase) was used to mimic a cytosolic source of redu cing equivalents. Oxygen pulses promote an acidification-alkalinizatio n proton cycle which is insensitive to antimycin and myxothiazol inhib itory effect, is stimulated by valinomycin, inhibited by trypsin-aprot inin complex, abolished by the protonophore carbonyl cyanide-p-trifluo romethoxy phenylhydrazone (FCCP), and is absent or at least inverted ( alkalinization-acidification cycle) in broken mitochondria. The oxidat ion of cytosolic substrates, mediated by the bi-trans-membrane electro n transport chain, does not involve endogenous cytochrome c and is ass ociated with a vectorial proton translocation from the inside to the o utside of the mitochondria. In the out --> in electron transport pathw ay the components involved appear to be cytosolic reduced substrates - -> NADH produced by cytosolic dehydrogenases activity --> NADH-cytochr ome b(5) oxidoreductase complex leaning out the external side of the e xternal membrane --> exogenous cytochrome c --> cytochrome oxidase of contact sites --> molecular oxygen. The possible components of the in --> out pathway are matrix respiratory substrates --> primary dehydrog enases of the matrix --> Complexes I, II, and III of the respiratory c hain present in the inner membrane --> NADH-cytochrome b(5) oxidoreduc tase system of the external membrane --> exogenous cytochrome c --> ad ditional cytosolic electron accepters or, alternatively, cytochrome ox idase of contact sites. The two pathways can be considered a bi-trans- membrane electron channeling system which, at the level of bridges set up by the contact points between the outer and the inner mitochondria l membrane, may represent a Link between the redox processes occurring inside with those present outside the mitochondrion. (C) 1995 Academi c Press, Inc.