ACTIVE UNFOLDING OF PRECURSOR PROTEINS DURING MITOCHONDRIAL PROTEIN IMPORT

Precursor proteins made in the cytoplasm must be in an unfolded confor mation during import into mitochondria, Some precursor proteins have t ightly folded domains but are imported faster than they unfold spontan eously, implying that mitochondria can unfold proteins, We measured th e import rates of artificial precursors containing presequences of var ying length fused to either mouse dihydrofolate reductase or bacterial barnase, and found that unfolding of a precursor at the mitochondrial surface is dramatically accelerated when its presequence is long enou gh to span both membranes and to interact with mhsp70 in the mitochond rial matrix, If the presequence is too short, import is slow but can b e strongly accelerated by urea-induced unfolding, suggesting that impo rt of these 'short' precursors is limited by spontaneous unfolding at the mitochondrial surface, With precursors that have sufficiently long presequences, unfolding by the inner membrane import machinery can be orders of magnitude faster than spontaneous unfolding, suggesting tha t mhsp70 can act as an ATP-driven force-generating motor during protei n import.