The Tim core complex defines the number of mitochondrial translocation contact sites and can hold arrested preproteins in the absence of matrix Hsp70-Tim44.
Preprotein import into mitochondria is mediated by translocases located in the outer and inner membranes (Tom and Tim) and a matrix Hsp70-Tim44 driving system. By blue native electrophoresis, we identify an approximately 90K complex with assembled Tim23 and Tim17 as the core of the inner membrane import site for presequence-containing preproteins. Preproteins spanning the two membranes link virtually all Tim core complexes with one in four Tom complexes in a stable 600K supercomplex. Neither mtHsp70 nor Tim44 are present in stoichiometric amounts in the 600K complex. Preproteins in transit stabilize the Tim core complex, preventing an exchange of subunits. Our studies define a central role for the Tim core complexes in mitochondrial protein import; they are not passive diffusion channels, but can stably interact with preproteins and determine the number of translocation contact sites. We propose the hypothesis that mtHsp70 functions in protein import not only by direct interaction with preproteins, but also by exerting a regulatory effect on the Tim channel.
Dekker
P J
PJ
Institut für Biochemie und Molekularbiologie, Universität Freiburg, Hermann-Herder-Str. 7, D-79104 Freiburg, Germany.
Martin
F
F
Maarse
A C
AC
Bömer
U
U
Müller
H
H
Guiard
B
B
Meijer
M
M
Rassow
J
J
Pfanner
N
N
D000595:Amino Acid Sequence; D001693:Biological Transport, Active; D002352:Carrier Proteins / Q000378:metabolism*; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism*; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D042844:L-Lactate Dehydrogenase (Cytochrome); D008565:Membrane Proteins / Q000378:metabolism*; D026901:Membrane Transport Proteins; D008928:Mitochondria / Q000378:metabolism*; D033681:Mitochondrial Membrane Transport Proteins; D000090622:Mitochondrial Precursor Protein Import Complex Proteins; D008969:Molecular Sequence Data; D011498:Protein Precursors / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012097:Repressor Proteins; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D013762:Tetrahydrofolate Dehydrogenase / Q000378:metabolism
D002352:Carrier Proteins; D018840:HSP70 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; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; C091191:TIM17 protein, S cerevisiae; C083081:TIM23 protein, S cerevisiae; C077016:TIM44 protein, S cerevisiae; D007770:L-Lactate Dehydrogenase; D042844:L-Lactate Dehydrogenase (Cytochrome); D013762:Tetrahydrofolate Dehydrogenase