THE ELECTROCHEMICAL-PROTON-GRADIENT-ACTIVATED STATES OF F0F1 ATPASE IN PLANT-MITOCHONDRIA AS REVEALED BY DETERGENTS

ATP hydrolysis, triggered by the addition of polyoxyethylene-9-lauryl ether (Lubrol) or lauryldimethylamine oxide (LDAO) to energized plant mitochondria was studied in some details. The membrane disruption was quasi-instantaneous (2-3 s) with both detergents, as shown by the decr ease of turbidity and the stopping of respiration. In pea leaf mitocho ndria, Lubrol triggered ATP hydrolysis in almost the same way as valin omycin plus nigericin, except that the activity was slightly stimulate d and became insensitive to carboxyatractyloside. This allowed investi gations of ATP hydrolysis without any interference of the ATP/ADP anti porter or the phosphate carrier. Lubrol did not prevent the ATPase fro m deactivating in pea leaf mitochondria, and did not trigger any ATP h ydrolysis in potato tuber mitochondria. At variance with Lubrol, LDAO changed the properties of the F0F1 ATPase. It made the enzyme oligomyc in insensitive and froze it in an activated state. The activity was al so 5-8-times stimulated in pea leaf mitochondria. Moreover, LDAO revea led an important ATP hydrolase activity when added to energized potato tuber mitochondria. Despite the specific effect of LDAO, the activity triggered by this detergent strongly depended on the energized state of the organelles before detergent addition. From this study, it is co ncluded that the electrochemical proton gradient is completely necessa ry to activate the F0F1-ATPase in intact plant mitochondria, as known in chloroplasts and suggested by some reports in animal mitochondria. Moreover, it is suggested that the main difference between the enzymes of pea leaf and potato tuber mitochondria is their rate of deactivati on after the collapse of the transmembrane electrochemical potential d ifference. Finally, when properly used, detergents appear to be a powe rful tool to probe the state of the ATPase in intact mitochondria, and maybe in more integrated systems.