Mitochondrial protein import: biochemical and genetic evidence for interaction of matrix hsp70 and the inner membrane protein MIM44.
The import of preproteins into mitochondria involves translocation of the polypeptide chains through putative channels in the outer and inner membranes. Preprotein-binding proteins are needed to drive the unidirectional translocation of the precursor polypeptides. Two of these preprotein-binding proteins are the peripheral inner membrane protein MIM44 and the matrix heat shock protein hsp70. We report here that MIM44 is mainly exposed on the matrix side, and a fraction of mt-hsp70 is reversibly bound to the inner membrane. Mt-hsp70 binds to MIM44 in a 1:1 ratio, suggesting that mt-hsp70 is localizing to the membrane via its interaction with MIM44. Formation of the complex requires a functional ATPase domain of mt-hsp70. Addition of Mg-ATP leads to dissociation of the complex. Overexpression of mt-hsp70 rescues the protein import defect of mutants in MIM44; conversely, overexpression of MIM44 rescues protein import defects of mt-hsp70 mutants. In addition, yeast strains with conditional mutations in both MIM44 and mt-hsp70 are barely viable, showing a synthetic growth defect compared to strains carrying single mutations. We propose that MIM44 and mt-hsp70 cooperate in translocation of preproteins. By binding to MIM44, mt-hsp70 is recruited at the protein import sites of the inner membrane, and preproteins arriving at MIM44 may be directly handed over to mt-hsp70.
Rassow
J
J
Biochemisches Institut, Universität Freiburg, Germany.
Maarse
A C
AC
Krainer
E
E
Kübrich
M
M
Müller
H
H
Meijer
M
M
Craig
E A
EA
Pfanner
N
N
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000595:Amino Acid Sequence; D001692:Biological Transport; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002451:Cell Compartmentation; D003432:Cross-Linking Reagents; D005656:Fungal Proteins / Q000235:genetics; D050956:HSP40 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism*; D006360:Heat-Shock Proteins / Q000235:genetics; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D026901:Membrane Transport Proteins; D008928:Mitochondria / Q000235:genetics / Q000378:metabolism*; D033681:Mitochondrial Membrane Transport Proteins; D000090622:Mitochondrial Precursor Protein Import Complex Proteins; D008969:Molecular Sequence Data; D009154:Mutation; D011233:Precipitin Tests; D011485:Protein Binding / Q000187:drug effects; D011498:Protein Precursors / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D013489:Suppression, Genetic
D002352:Carrier Proteins; D003432:Cross-Linking Reagents; D005656:Fungal Proteins; D050956:HSP40 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D033681:Mitochondrial Membrane Transport Proteins; D000090622:Mitochondrial Precursor Protein Import Complex Proteins; D011498:Protein Precursors; C074215:SEC63 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C077016:TIM44 protein, S cerevisiae; D000255:Adenosine Triphosphate