Mitochondrial protein import: molecular basis of the ATP-dependent interaction of MtHsp70 with Tim44.
Protein translocation across the mitochondrial inner membrane is driven by cycles of binding and release of mitochondrial heat shock protein 70 (mtHsp70) in the matrix. The peripheral inner membrane protein, Tim44, recruits mtHsp70 in an ATP-dependent manner to the import sites. We show that DnaK, the closely related Hsp70 of Escherichia coli, when targeted to the matrix of yeast mitochondria, interacts in a specific manner with Tim44. The interaction is, however, not regulated by ATP, and DnaK cannot support protein translocation. We used truncated mtHsp70s and chimeric proteins consisting of segments of mtHsp70 and DnaK to analyze which portions of mtHsp70 bind and functionally interact with Tim44. We show that Tim44 interacts with the beta-stranded core of the peptide binding domain of mtHsp70 and of DnaK. The alpha-helices A and B of the peptide binding domain of mtHsp70 are required to transmit the nucleotide state of the ATPase domain to the peptide binding domain. Tim44, by interacting in this way with the peptide binding domain, is proposed to coordinate the binding of mtHsp70 to the incoming preprotein and the subsequent release of the mtHsp70-preprotein complex from the TIM23 complex, the translocase of the inner membrane.
Moro
Fernando
F
Institut für Physiologische Chemie der Universität München, Butenandtstrasse 5, 81377 Munich, Germany.
Okamoto
Koji
K
Donzeau
Mariel
M
Neupert
Walter
W
Brunner
Michael
M
D000255:Adenosine Triphosphate / Q000378:metabolism*; D000595:Amino Acid Sequence; D000252:Calcium-Transporting ATPases / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism; D029968:Escherichia coli Proteins; D005656:Fungal Proteins / Q000378:metabolism; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism*; D006360:Heat-Shock Proteins; D008565:Membrane Proteins / Q000378:metabolism*; D026901:Membrane Transport Proteins; D008928:Mitochondria / Q000378:metabolism*; D033681:Mitochondrial Membrane Transport Proteins; D008958:Models, Molecular; D018832:Molecular Chaperones / Q000378:metabolism; D008969:Molecular Sequence Data; D011233:Precipitin Tests; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D013997:Time Factors
D002352:Carrier Proteins; D029968:Escherichia coli Proteins; D005656:Fungal Proteins; D018840:HSP70 Heat-Shock Proteins; D006360:Heat-Shock Proteins; C086039:MGE1 protein, S cerevisiae; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D033681:Mitochondrial Membrane Transport Proteins; D018832:Molecular Chaperones; D011993:Recombinant Fusion Proteins; C101767:SSC1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D000255:Adenosine Triphosphate; C063842:dnaK protein, E coli; D000252:Calcium-Transporting ATPases