KH-17, a simplified derivative of bongkrekic acid, weakly inhibits the mitochondrial ADP/ATP carrier from both sides of the inner mitochondrial membrane

Two natural products, bongkrekic acid and carboxyatractyloside, are known to specifically inhibit the mitochondrial ADP/ATP carrier from its matrix side and cytosolic side, respectively, in concentration ranges of 10(-6) M. In the present study, we investigated the manner of action of a synthetic bongkrekic acid derivative, KH-17, lacking three methyl groups, one methoxy group, and five internal double bonds, on the mitochondrial ADP/ATP carrier. At slightly acidic pH, KH-17 inhibited mitochondrial [H-3]ADP uptake, but its inhibitory action was about 10 times weaker than that of its parental compound, bongkrekic acid. The main site of action of KH-17 was confirmed as the matrix side of the ADP/ATP carrier by experiments using submitochondrial particles, which have an inside-out orientation of the inner mitochondrial membrane. However, when we added KH-17 to mitochondria at neutral pH, it had a weak inhibitory effect on [H-3]ADP uptake, and its inhibitory strength was similar to that of bongkrekic acid. These results indicated that KH-17 weakly inhibits the ADP/ATP carrier not only from the matrix side but also from the cytosolic side. To ascertain whether this interpretation was correct, we examined the effects of KH-17 and carboxyatractyloside on mitochondrial [H-3]ADP uptake at two [H-3]ADP concentrations. We found that both KH-17 and carboxyatractyloside showed a stronger inhibitory effect at the lower [H-3]ADP concentration. Therefore, we concluded that the bongkrekic acid derivative, KH-17, weakly inhibits the mitochondrial ADP/ATP carrier from both sides of the inner mitochondrial membrane. These results suggested that the elimination of three methyl groups, one methoxy group, and five internal double bonds present in bongkrekic acid altered its manner of action towards the mitochondrial ADP/ATP carrier. Our data will help to improve our understanding of the interaction between bongkrekic acid and the mitochondrial ADP/ATP carrier.