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Title: Two alternative binding mechanisms connect the protein translocation Sec71-Sec72 complex with heat shock proteins

Journal Article · · Journal of Biological Chemistry
 [1];  [2];  [2];  [1]
  1. Harvard Medical School, Boston, MA (United States). Howard Hughes Medical Inst. Dept. of Cell Biology
  2. Univ. of Massachusetts Medical School, Worcester, MA (United States). Dept. of Biochemistry and Molecular Pharmacology
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The biosynthesis of many eukaryotic proteins requires accurate targeting to and translocation across the endoplasmic reticulum membrane. Post-translational protein translocation in yeast requires both the Sec61 translocation channel, and a complex of four additional proteins: Sec63, Sec62, Sec71, and Sec72. The structure and function of these proteins are largely unknown. This pathway also requires the cytosolic Hsp70 protein Ssa1, but whether Ssa1 associates with the translocation machinery to target protein substrates to the membrane is unclear. Here, we use a combined structural and biochemical approach to explore the role of Sec71-Sec72 subcomplex in post-translational protein translocation. To this end, we report a crystal structure of the Sec71-Sec72 complex, which revealed that Sec72 contains a tetratricopeptide repeat (TPR) domain that is anchored to the endoplasmic reticulum membrane by Sec71. We also determined the crystal structure of this TPR domain with a C-terminal peptide derived from Ssa1, which suggests how Sec72 interacts with full-length Ssa1. Surprisingly, Ssb1, a cytoplasmic Hsp70 that binds ribosome-associated nascent polypeptide chains, also binds to the TPR domain of Sec72, even though it lacks the TPR-binding C-terminal residues of Ssa1. We demonstrate that Ssb1 binds through its ATPase domain to the TPR domain, an interaction that leads to inhibition of nucleotide exchange. Taken together, our results suggest that translocation substrates can be recruited to the Sec71-Sec72 complex either post-translationally through Ssa1 or co-translationally through Ssb1.

Research Organization:
Argonne National Laboratory (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
Sponsoring Organization:
USDOE Office of Science (SC)
OSTI ID:
1372257
Journal Information:
Journal of Biological Chemistry, Journal Name: Journal of Biological Chemistry Journal Issue: 19 Vol. 292; ISSN 0021-9258
Publisher:
American Society for Biochemistry and Molecular BiologyCopyright Statement
Country of Publication:
United States
Language:
ENGLISH

References (48)

Secretion in yeast: translocation and glycosylation of prepro-α-factor in vitro can occur via an ATP-dependent post-translational mechanism journal May 1986
The signal recognition particle in S. cerevisiae journal October 1991
The translation machinery and 70 kd heat shock protein cooperate in protein synthesis journal October 1992
Posttranslational protein transport in yeast reconstituted with a purified complex of Sec proteins and Kar2p journal May 1995
Preparation of selenomethionyl proteins for phase determination book January 1997
BiP Acts as a Molecular Ratchet during Posttranslational Transport of Prepro-α Factor across the ER Membrane journal May 1999
Structure of TPR Domain–Peptide Complexes journal April 2000
Molecular Chaperones Hsp90 and Hsp70 Deliver Preproteins to the Mitochondrial Import Receptor Tom70 journal January 2003
J Proteins Catalytically Activate Hsp70 Molecules to Trap a Wide Range of Peptide Sequences journal November 1998
The ribosome-bound Hsp70 homolog Ssb of Saccharomyces cerevisiae journal June 2010
Protein import machineries of peroxisomes journal March 2011
Structural Basis for the Cooperation of Hsp70 and Hsp110 Chaperones in Protein Folding journal June 2008
The Cotranslational Function of Ribosome-Associated Hsp70 in Eukaryotic Protein Homeostasis journal January 2013
The Myosin Chaperone UNC-45 Is Organized in Tandem Modules to Support Myofilament Formation in C. elegans journal January 2013
A Network of Cytosolic Factors Targets SRP-Independent Proteins to the Endoplasmic Reticulum journal February 2013
Inference of Macromolecular Assemblies from Crystalline State journal September 2007
A subfamily of stress proteins facilitates translocation of secretory and mitochondrial precursor polypeptides journal April 1988
70K heat shock related proteins stimulate protein translocation into microsomes journal April 1988
Assembly of yeast Sec proteins involved in translocation into the endoplasmic reticulum into a membrane-bound multisubunit complex journal February 1991
Structure and function of Hip, an attenuator of the Hsp70 chaperone cycle journal June 2013
Crystal structure of yeast Sis1 peptide-binding fragment and Hsp70 Ssa1 C-terminal complex journal August 2006
Molecular Chaperone Hsp70/Hsp90 Prepares the Mitochondrial Outer Membrane Translocon Receptor Tom71 for Preprotein Loading journal August 2009
Structural Studies and the Assembly of the Heptameric Post-translational Translocon Complex journal January 2011
A Structural Model of the Sgt2 Protein and Its Interactions with Chaperones and the Get4/Get5 Complex journal September 2011
Secretory protein translocation in a yeast cell-free system can occur posttranslationally and requires ATP hydrolysis. journal May 1986
Prepro-carboxypeptidase Y and a truncated form of pre-invertase, but not full-length pre-invertase, can be posttranslationally translocated across microsomal vesicle membranes from Saccharomyces cerevisiae. journal April 1988
Multiple genes are required for proper insertion of secretory proteins into the endoplasmic reticulum in yeast. journal December 1989
Sec72p contributes to the selective recognition of signal peptides by the secretory polypeptide translocation complex. journal August 1994
Signal sequences specify the targeting route to the endoplasmic reticulum membrane. journal July 1996
Spontaneous Release of Cytosolic Proteins from Posttranslational Substrates before Their Transport into the Endoplasmic Reticulum journal October 2000
Identification of cytoplasmic residues of Sec61p involved in ribosome binding and cotranslational translocation journal January 2005
Structural and functional characterization of Sec66p, a new subunit of the polypeptide translocation apparatus in the yeast endoplasmic reticulum. journal September 1993
DaliLite workbench for protein structure comparison journal June 2000
The Tetratricopeptide Repeats of Receptors Involved in Protein Translocation across Membranes journal June 2007
Vectors for co-expression of an unrestricted number of proteins journal March 2007
Dali server: conservation mapping in 3D journal May 2010
Protein Translocation across the Rough Endoplasmic Reticulum journal December 2012
Phaser crystallographic software journal July 2007
MolProbity : all-atom structure validation for macromolecular crystallography journal December 2009
XDS journal January 2010
PHENIX: a comprehensive Python-based system for macromolecular structure solution journal January 2010
Features and development of Coot journal March 2010
Principles of ER cotranslational translocation revealed by proximity-specific ribosome profiling journal November 2014
Structural and functional dissection of Sec62p, a membrane-bound component of the yeast endoplasmic reticulum protein import machinery. journal November 1990
Topology and functional domains of Sec63p, an endoplasmic reticulum membrane protein required for secretory protein translocation. journal July 1992
Functional interaction of cytosolic hsp70 and a DnaJ-related protein, Ydj1p, in protein translocation in vivo journal August 1996
Mechanisms of Sec61/SecY-Mediated Protein Translocation Across Membranes journal June 2012
Ligand Recognition by the TPR Domain of the Import Factor Toc64 from Arabidopsis thaliana journal December 2013

Cited By (11)

Post-Translational Protein Transport by the Sec Complex journal June 2019
MANF antagonizes nucleotide exchange by the endoplasmic reticulum chaperone BiP text January 2019
MANF antagonizes nucleotide exchange by the endoplasmic reticulum chaperone BiP journal February 2019
Structure of the post-translational protein translocation machinery of the ER membrane journal December 2018
Binding properties of the quaternary assembly protein SPAG1 journal June 2019
Conserved motifs on the cytoplasmic face of the protein translocation channel are critical for the transition between resting and active conformations journal August 2018
Guiding tail-anchored membrane proteins to the endoplasmic reticulum in a chaperone cascade journal October 2019
Structure of the posttranslational Sec protein-translocation channel complex from yeast journal December 2018
Targeting and translocation of proteins to the endoplasmic reticulum at a glance journal December 2017
A clearer picture of the ER translocon complex journal February 2020
Substrate relay in an Hsp70‐cochaperone cascade safeguards tail‐anchored membrane protein targeting journal July 2018

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Related Subjects

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY
59 BASIC BIOLOGICAL SCIENCES
70 kilodalton heat shock protein (Hsp70)
Sec71
Sec72
Ssa1
Ssb1
TPR
endoplasmic reticulum (ER)
protein structure
protein translocation
structure-function