The role of the Tim8p-Tim13p complex in a conserved import pathway for mitochondrial polytopic inner membrane proteins.
Tim23p is imported via the TIM (translocase of inner membrane)22 pathway for mitochondrial inner membrane proteins. In contrast to precursors with an NH2-terminal targeting presequence that are imported in a linear NH2-terminal manner, we show that Tim23p crosses the outer membrane as a loop before inserting into the inner membrane. The Tim8p-Tim13p complex facilitates translocation across the intermembrane space by binding to the membrane spanning domains as shown by Tim23p peptide scans with the purified Tim8p-Tim13p complex and crosslinking studies with Tim23p fusion constructs. The interaction between Tim23p and the Tim8p-Tim13p complex is not dependent on zinc, and the purified Tim8p-Tim13p complex does not coordinate zinc in the conserved twin CX3C motif. Instead, the cysteine residues seemingly form intramolecular disulfide linkages. Given that proteins of the mitochondrial carrier family also pass through the TOM (translocase of outer membrane) complex as a loop, our study suggests that this translocation mechanism may be conserved. Thus, polytopic inner membrane proteins, which lack an NH2-terminal targeting sequence, pass through the TOM complex as a loop followed by binding of the small Tim proteins to the hydrophobic membrane spanning domains.
Curran
Sean P
SP
Department of Chemistry and Biochemistry, University of California Los Angeles, 607 Charles Young Drive East, Los Angeles, CA 90095, USA.
Leuenberger
Danielle
D
Schmidt
Einhard
E
Koehler
Carla M
CM
D002352:Carrier Proteins / Q000378:metabolism* / Q000502:physiology*; D002942:Circular Dichroism; D004220:Disulfides / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D007425:Intracellular Membranes / Q000378:metabolism; D046911:Macromolecular Substances; D008565:Membrane Proteins / Q000378:metabolism; D026901:Membrane Transport Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism*; D033681:Mitochondrial Membrane Transport Proteins; D000090622:Mitochondrial Precursor Protein Import Complex Proteins; D024101:Mitochondrial Proteins / Q000378:metabolism*; D008970:Molecular Weight; D021381:Protein Transport; D012441:Saccharomyces cerevisiae / Q000737:chemistry / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism; D015032:Zinc / Q000032:analysis
D002352:Carrier Proteins; D004220:Disulfides; D046911:Macromolecular Substances; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D033681:Mitochondrial Membrane Transport Proteins; D000090622:Mitochondrial Precursor Protein Import Complex Proteins; D024101:Mitochondrial Proteins; D029701:Saccharomyces cerevisiae Proteins; C403093:TIM13 protein, S cerevisiae; C083081:TIM23 protein, S cerevisiae; C485737:TIMM13 protein, human; C118122:TIMM8A protein, human; D015032:Zinc