Tim9p, an essential partner subunit of Tim10p for the import of mitochondrial carrier proteins

Tim10p, a protein of the yeast mitochondrial intermembrane space, was shown previously to be essential for the import of multispanning carrier protein s from the cytoplasm into the inner membrane. We now identify Tim9p, anothe r essential component of this import pathway. Most of Tim9p is associated w ith Tim10p in a soluble 70 kDa complex. Tim9p and Tim10p copurify in succes sive chromatographic fractionations and co-immunoprecipitated with each oth er, Tim9p can be cross-linked to a partly translocated carrier protein, A s mall fraction of Tim9p is bound to the outer face of the inner membrane in a 300 kDa complex whose other subunits include Tim54p, Tim22p, Tim12p and T im10p, The sequence of Tim9p is 25% identical to that of Tim10p and Tim12p, A Ser(67)-->Cys(67) mutation in Tim9p suppresses the temperature-sensitive growth defect of tim10-1 and tim12-1 mutants. Tim9p is a new subunit of th e TIM machinery that guides hydrophobic inner membrane proteins across the aqueous intermembrane space.