The mitochondrial aspartate/glutamate and ADP/ATP carrier switch from obligate counterexchange to unidirectional transport after modification by SH-reagents.

The influence of various SH-reagents on the aspartate/glutamate carrier was investigated in the reconstituted system. When liposomes carrying partially purified carrier protein were treated with 5,5'-dithiobis(2-nitrobenzoic acid) or N-ethylmaleimide, antiport activity was strongly reduced. Several mercury compounds exerted a dual effect. They completely blocked the antiport and, in addition, induced an efflux pathway for internal aspartate. The maximum rate of this unidirectional flux was comparable to the original antiport activity. Induction of efflux always was coupled to inhibition of antiport. Efflux was neither due to unspecific leakage of proteoliposomes nor to a possible contamination by porin, but depended on active carrier protein, as elucidated by the sensitivity to proteinases and protein-modifying reagents. Besides efflux of aspartate, HgCl2 and mersalyl also induced a slow efflux of ATP from liposomes carrying coreconstituted aspartate/glutamate and ADP/ATP carrier. The two efflux activities could be discriminated taking advantage of the differential effectiveness of several inhibitors and proteinases. Although basic carrier properties were changed by the applied mercurials (Dierks, T., Salentin, A. and Krämer, R. (1990) Biochim. Biophys. Acta 1028, 281), aspartate and ATP efflux could clearly be correlated with the aspartate/glutamate and the ADP/ATP carrier, respectively. When purifying these two translocators the respective efflux activity copurified with the antiporter, thus elucidating that the two different transport functions are mediated by the same protein. These results argue for a participation of the aspartate/glutamate and the ADP/ATP carrier in the generally observed increase of mitochondrial permeability after treatment with SH-reagents.