IMPORT OF SUBUNIT-VII OF THE CYTOCHROME BC(1) COMPLEX INTO YEAST MITOCHONDRIA

The import of subunit VII, the so-called ubiquinol binding protein of the cytochrome bc(1) complex of yeast mitochondria, has been investiga ted by subcloning the gene for subunit VII into the expression vector pSP64. The precursor to subunit VII synthesized in vitro by a transcri ption/translation system had the same molecular weight as the mature f orm of this protein present in mitochondria, confirming earlier report s that subunit VII does not contain a cleavable presequence. The subun it VII precursor was imported efficiently into a protease-resistant co mpartment of isolated yeast mitochondria in a reaction dependent upon a membrane potential and the presence of ATP both in the mitochondrial matrix and in the extramitochondrial space. After import, the radiola beled subunit VII was bound to the mitochondrial inner membrane throug h protein-protein interactions and could not be detected in the matrix fraction or in the intermembrane space. Subunit VII was also imported into mitoplasts, where the protein was associated with the inner memb rane facing the matrix side of the membrane. By contrast, after import into mitochondria obtained from yeast cells lacking cytochrome b, sub unit VII was localized in the supernatant fraction, suggesting that th e presence of cytochrome b may be required for the association of subu nit VII with the inner mitochondrial membrane. (C) 1995 Academic Press , Inc.