title: The Tom40 assembly process probed using the attachment of different intramitochondrial sorting signals.
abstract: The TOM40 complex is a protein translocator in the mitochondrial outer membrane and consists of several different subunits. Among them, Tom40 is a central subunit that constitutes a protein-conducting channel by forming a β-barrel structure. To probe the nature of the assembly process of Tom40 in the outer membrane, we attached various mitochondrial presequences to Tom40 that possess sorting information for the intermembrane space (IMS), inner membrane, and matrix and would compete with the inherent Tom40 assembly process. We analyzed the mitochondrial import of those fusion proteins in vitro. Tom40 crossed the outer membrane and/or inner membrane even in the presence of various sorting signals. N-terminal anchorage of the attached presequence to the inner membrane did not prevent Tom40 from associating with the TOB/SAM complex, although it impaired its efficient release from the TOB complex in vitro but not in vivo. The IMS or matrix-targeting presequence attached to Tom40 was effective in substituting for the requirement for small Tim proteins in the IMS for the translocation of Tom40 across the outer membrane. These results provide insight into the mechanism responsible for the precise delivery of β-barrel proteins to the outer mitochondrial membrane.
authors.affiliation: Department of Chemistry, Graduate School of Science, Nagoya University, Chikusa-ku, Nagoya 464-8602, Japan.
authors.forename: Takuya
authors.initials: T
authors.lastname: Shiota
authors.forename: Miyuki
authors.initials: M
authors.lastname: Maruyama
authors.forename: Mami
authors.initials: M
authors.lastname: Miura
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Tamura
authors.forename: Koji
authors.initials: K
authors.lastname: Yamano
authors.forename: Masatoshi
authors.initials: M
authors.lastname: Esaki
authors.forename: Toshiya
authors.initials: T
authors.lastname: Endo
mesh_terms: D053078:Membrane Potential, Mitochondrial; D008928:Mitochondria / Q000378:metabolism* / Q000648:ultrastructure; D033681:Mitochondrial Membrane Transport Proteins / Q000737:chemistry* / Q000378:metabolism* / Q000648:ultrastructure; D051336:Mitochondrial Membranes / Q000378:metabolism / Q000648:ultrastructure; D008954:Models, Biological; D011499:Protein Processing, Post-Translational; D021382:Protein Sorting Signals; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism / Q000648:ultrastructure; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000378:metabolism* / Q000648:ultrastructure
chemical_list: D033681:Mitochondrial Membrane Transport Proteins; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; C066555:Tom40 protein, S cerevisiae