Many nuclear-encoded mitochondrial proteins, which are synthesized as precursors bearing an amino-terminal presequence, have to move across one or two mitochondrial mambranes to reach their destinations. Our physicochemical studys on a model precursor protein, pCOX IV-DHFR, suggest that the initial binding of this protein to the mitochondrial outer surface requires lipid-mediated partial unfolding of the polypeptide chain. Then the presequence of the precursor protein moves across the outer and inner membranes via contact sites in a membrane potential-dependent manner, and the traslocation of the mature part across the two membranes follows.