title: Structural studies and the assembly of the heptameric post-translational translocon complex.
abstract: In Saccharomyces cerevisiae, some of the nascent chains can be post-translationally translocated into the endoplasmic reticulum through the heptameric post-translational translocon complex (post-translocon). This membrane-protein complex is composed of the protein-conducting channel and the tetrameric Sec62/63 complex. The Sec62/63 complex plays crucial roles in targeting of the signal recognition particle-independent protein substrate to the protein-conducting channel and in assembly of the post-translocon. Although the molecular mechanism of the post-translational translocation process has been well established, the structure of the post-translocon and how the channel and the Sec62/63 complex form the heptameric complex are largely uncharacterized. Here, we report a 20-Å resolution cryo-electron microscopy structure of the post-translocon. The purified post-translocon was found to have a mass of 287 kDa, which is consistent with the unit stoichiometry of the seven subunits as determined by a cysteine labeling experiment. We demonstrated that Triton X-100 dissociated the heptameric complex into three subcomplexes identified as the trimeric translocon Sec61-Sbh1-Sss1, the Sec63-Sec71-Sec72 trimer, and the heterotetramer Sec62-Sec63-Sec71-Sec72, respectively. Additionally, a role of the sixth cytosolic loop of Sec61 in assembly of the post-translocon was demonstrated. Mutations of conserved, positively charged amino acid residues in the loop caused decreased formation of the post-translocon. These studies provide the first architectural description of the yeast post-translocon.
authors.affiliation: Department of Biochemistry and Cell Biology, Stony Brook University, Stony Brook, New York 11794, USA.
authors.forename: Yoichiro
authors.initials: Y
authors.lastname: Harada
authors.forename: Hua
authors.initials: H
authors.lastname: Li
authors.forename: Joseph S
authors.initials: JS
authors.lastname: Wall
authors.forename: Huilin
authors.initials: H
authors.lastname: Li
authors.forename: William J
authors.initials: WJ
authors.lastname: Lennarz
mesh_terms: D020285:Cryoelectron Microscopy; D004721:Endoplasmic Reticulum / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D026901:Membrane Transport Proteins; D046912:Multiprotein Complexes / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D009154:Mutation; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D021381:Protein Transport / Q000502:physiology; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D013329:Structure-Activity Relationship
chemical_list: D026901:Membrane Transport Proteins; D046912:Multiprotein Complexes; D029701:Saccharomyces cerevisiae Proteins
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