IMPORTANCE OF LOOPS OF MITOCHONDRIAL ADP/ATP CARRIER FOR ITS TRANSPORT ACTIVITY DEDUCED FROM REACTIVITIES OF ITS CYSTEINE RESIDUES WITH THE SULFHYDRYL REAGENT EOSIN-5-MALEIMIDE

The effects of various compounds such as the transport substrate ADP and the transport inhibitors carboxyatractyloside (CATR) and bongkrekic acid (BKA) on the labeling of cysteine residues in the ADP/ ATP carrier of bovine heart submitochondrial particles by the SH reagent eosin-5-maleimide (EMA) were studied. Of the four cysteine residues in the carrier, the labeling of Cys(159) by EMA progressed predominantly and rapidly, and those of Cys(56),,d Cys(256) moderately, but Cys(128) was not labeled, as we reported previously [Majima, E., et al. (1993) J. Biol. Chem. 268, 22181-22187]. ADP inhibited the labelings of Cys(56), Cys(159) and Cys(256) by EMA. BKA markedly inhibited the labeling of Cys(159) by EMA, and also the labeling of. Cys(256), but did not affect the labeling of Cys(56) suggesting that it binds from the matrix side to a region close to Cys(159) in the second loop facing the matrix space. CATR completely inhibited the labeling by EMA when added on the cytosolic side, but had no effect when added on the matrix side. From these results, the conformational changes of the carrier induced by CATR, BKA, and ADP are discussed. Furthermore, a mechanism of adenine nucleotide transport through the ADP/ATP carrier in association with change in its conformation is proposed.