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Title: Structural determinants for protein unfolding and translocation by the Hsp104 protein disaggregase

Abstract

Here, the ring-forming Hsp104 ATPase cooperates with Hsp70 and Hsp40 molecular chaperones to rescue stress-damaged proteins from both amorphous and amyloid-forming aggregates. The ability to do so relies upon pore loops present in the first ATP-binding domain (AAA-1; loop-1 and loop-2) and in the second ATP-binding domain (AAA-2; loop-3) of Hsp104, which face the protein translocating channel and couple ATP-driven changes in pore loop conformation to substrate translocation. A hallmark of loop-1 and loop-3 is an invariable and mutational sensitive aromatic amino acid (Tyr257 and Tyr662) involved in substrate binding. However, the role of conserved aliphatic residues (Lys256, Lys258, and Val663) flanking the pore loop tyrosines, and the function of loop-2 in protein disaggregation has not been investigated. Here we present the crystal structure of an N-terminal fragment of Saccharomyces cerevisiae Hsp104 exhibiting molecular interactions involving both AAA-1 pore loops, which resemble contacts with bound substrate. Corroborated by biochemical experiments and functional studies in yeast, we show that aliphatic residues flanking Tyr257 and Tyr662 are equally important for substrate interaction, and abolish Hsp104 function when mutated to glycine. Unexpectedly, we find that loop-2 is sensitive to aspartate substitutions that impair Hsp104 function and abolish protein disaggregation when loop-2 is replacedmore » by four aspartate residues. Our observations suggest that Hsp104 pore loops have non-overlapping functions in protein disaggregation and together coordinate substrate binding, unfolding, and translocation through the Hsp104 hexamer.« less

Authors:
 [1];  [1];  [1];  [2];  [1];  [1]
  1. Baylor College of Medicine, Houston, TX (United States)
  2. Argonne National Lab. (ANL), Argonne, IL (United States)
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States)
Sponsoring Org.:
National Institutes of Health (NIH); USDOE Office of Science (SC), Biological and Environmental Research (BER) (SC-23)
OSTI Identifier:
1461438
Grant/Contract Number:  
AC02-06CH11357
Resource Type:
Accepted Manuscript
Journal Name:
Bioscience Reports
Additional Journal Information:
Journal Volume: 37; Journal Issue: 6; Journal ID: ISSN 0144-8463
Publisher:
Portland Press - Biochemical Society
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES

Citation Formats

Lee, Jungsoon, Sung, Nuri, Yeo, Lythou, Chang, Changsoo, Lee, Sukyeong, and Tsai, Francis T. F. Structural determinants for protein unfolding and translocation by the Hsp104 protein disaggregase. United States: N. p., 2017. Web. doi:10.1042/BSR20171399.
Lee, Jungsoon, Sung, Nuri, Yeo, Lythou, Chang, Changsoo, Lee, Sukyeong, & Tsai, Francis T. F. Structural determinants for protein unfolding and translocation by the Hsp104 protein disaggregase. United States. doi:10.1042/BSR20171399.
Lee, Jungsoon, Sung, Nuri, Yeo, Lythou, Chang, Changsoo, Lee, Sukyeong, and Tsai, Francis T. F. Fri . "Structural determinants for protein unfolding and translocation by the Hsp104 protein disaggregase". United States. doi:10.1042/BSR20171399. https://www.osti.gov/servlets/purl/1461438.
@article{osti_1461438,
title = {Structural determinants for protein unfolding and translocation by the Hsp104 protein disaggregase},
author = {Lee, Jungsoon and Sung, Nuri and Yeo, Lythou and Chang, Changsoo and Lee, Sukyeong and Tsai, Francis T. F.},
abstractNote = {Here, the ring-forming Hsp104 ATPase cooperates with Hsp70 and Hsp40 molecular chaperones to rescue stress-damaged proteins from both amorphous and amyloid-forming aggregates. The ability to do so relies upon pore loops present in the first ATP-binding domain (AAA-1; loop-1 and loop-2) and in the second ATP-binding domain (AAA-2; loop-3) of Hsp104, which face the protein translocating channel and couple ATP-driven changes in pore loop conformation to substrate translocation. A hallmark of loop-1 and loop-3 is an invariable and mutational sensitive aromatic amino acid (Tyr257 and Tyr662) involved in substrate binding. However, the role of conserved aliphatic residues (Lys256, Lys258, and Val663) flanking the pore loop tyrosines, and the function of loop-2 in protein disaggregation has not been investigated. Here we present the crystal structure of an N-terminal fragment of Saccharomyces cerevisiae Hsp104 exhibiting molecular interactions involving both AAA-1 pore loops, which resemble contacts with bound substrate. Corroborated by biochemical experiments and functional studies in yeast, we show that aliphatic residues flanking Tyr257 and Tyr662 are equally important for substrate interaction, and abolish Hsp104 function when mutated to glycine. Unexpectedly, we find that loop-2 is sensitive to aspartate substitutions that impair Hsp104 function and abolish protein disaggregation when loop-2 is replaced by four aspartate residues. Our observations suggest that Hsp104 pore loops have non-overlapping functions in protein disaggregation and together coordinate substrate binding, unfolding, and translocation through the Hsp104 hexamer.},
doi = {10.1042/BSR20171399},
journal = {Bioscience Reports},
number = 6,
volume = 37,
place = {United States},
year = {2017},
month = {12}
}

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Works referenced in this record:

Functional analysis of conserved cis- and trans-elements in the Hsp104 protein disaggregating machine
journal, August 2012

  • Biter, Amadeo B.; Lee, Jungsoon; Sung, Nuri
  • Journal of Structural Biology, Vol. 179, Issue 2
  • DOI: 10.1016/j.jsb.2012.05.007

Coordinated Hsp110 and Hsp104 Activities Power Protein Disaggregation in Saccharomyces cerevisiae
journal, March 2017

  • Kaimal, Jayasankar Mohanakrishnan; Kandasamy, Ganapathi; Gasser, Fabian
  • Molecular and Cellular Biology, Vol. 37, Issue 11
  • DOI: 10.1128/MCB.00027-17

Hsp70 targets Hsp100 chaperones to substrates for protein disaggregation and prion fragmentation
journal, August 2012

  • Winkler, Juliane; Tyedmers, Jens; Bukau, Bernd
  • The Journal of Cell Biology, Vol. 198, Issue 3
  • DOI: 10.1083/jcb.201201074

Escherichia coli ClpB is a non-processive polypeptide translocase
journal, August 2015

  • Li, Tao; Weaver, Clarissa L.; Lin, Jiabei
  • Biochemical Journal, Vol. 470, Issue 1
  • DOI: 10.1042/BJ20141457

Loops in the Central Channel of ClpA Chaperone Mediate Protein Binding, Unfolding, and Translocation
journal, July 2005


AAA+ Proteases: ATP-Fueled Machines of Protein Destruction
journal, July 2011


Role of the GYVG Pore Motif of HslU ATPase in Protein Unfolding and Translocation for Degradation by HslV Peptidase
journal, April 2005

  • Park, Eunyong; Rho, Young Min; Koh, Ohn-jo
  • Journal of Biological Chemistry, Vol. 280, Issue 24
  • DOI: 10.1074/jbc.M500035200

Ratchet-like polypeptide translocation mechanism of the AAA+ disaggregase Hsp104
journal, June 2017


Protein rescue from aggregates by powerful molecular chaperone machines
journal, September 2013

  • Doyle, Shannon M.; Genest, Olivier; Wickner, Sue
  • Nature Reviews Molecular Cell Biology, Vol. 14, Issue 10
  • DOI: 10.1038/nrm3660

Overlapping and Specific Functions of the Hsp104 N Domain Define Its Role in Protein Disaggregation
journal, September 2017


Inorganic phosphate assay with malachite green: An improvement and evaluation
journal, December 1982


In Vivo Monitoring of the Prion Replication Cycle Reveals a Critical Role for Sis1 in Delivering Substrates to Hsp104
journal, November 2008


Species-specific collaboration of heat shock proteins (Hsp) 70 and 100 in thermotolerance and protein disaggregation
journal, April 2011

  • Miot, M.; Reidy, M.; Doyle, S. M.
  • Proceedings of the National Academy of Sciences, Vol. 108, Issue 17
  • DOI: 10.1073/pnas.1102828108

Quantitative method for the assignment of hinge and shear mechanism in protein domain movements
journal, July 2014


Insights into Hsp70 Chaperone Activity from a Crystal Structure of the Yeast Hsp110 Sse1
journal, October 2007


Hsp104, Hsp70, and Hsp40
journal, July 1998


Structures of Asymmetric ClpX Hexamers Reveal Nucleotide-Dependent Motions in a AAA+ Protein-Unfolding Machine
journal, November 2009


Substrate recognition by the AAA+ chaperone ClpB
journal, June 2004

  • Schlieker, Christian; Weibezahn, Jimena; Patzelt, Holger
  • Nature Structural & Molecular Biology, Vol. 11, Issue 7
  • DOI: 10.1038/nsmb787

Thermotolerance Requires Refolding of Aggregated Proteins by Substrate Translocation through the Central Pore of ClpB
journal, November 2004


Heat shock protein (Hsp) 70 is an activator of the Hsp104 motor
journal, May 2013

  • Lee, J.; Kim, J. -H.; Biter, A. B.
  • Proceedings of the National Academy of Sciences, Vol. 110, Issue 21
  • DOI: 10.1073/pnas.1217988110

Crystal Structure of E.coli Hsp100 ClpB Nucleotide-binding Domain 1 (NBD1) and Mechanistic Studies on ClpB ATPase Activity
journal, May 2002


Structural basis for the disaggregase activity and regulation of Hsp104
journal, November 2016

  • Heuck, Alexander; Schitter-Sollner, Sonja; Suskiewicz, Marcin Józef
  • eLife, Vol. 5
  • DOI: 10.7554/eLife.21516

The M-Domain Controls Hsp104 Protein Remodeling Activity in an Hsp70/Hsp40-Dependent Manner
journal, September 2010


Mutational studies on HslU and its docking mode with HslV
journal, December 2000

  • Song, H. K.; Hartmann, C.; Ramachandran, R.
  • Proceedings of the National Academy of Sciences, Vol. 97, Issue 26
  • DOI: 10.1073/pnas.250491797

Mechanistic and Structural Insights into the Prion-Disaggregase Activity of Hsp104
journal, May 2016


Crystal and Solution Structures of an HslUV Protease–Chaperone Complex
journal, November 2000


Peptide and Protein Binding in the Axial Channel of Hsp104: INSIGHTS INTO THE MECHANISM OF PROTEIN UNFOLDING
journal, August 2008

  • Lum, Ronnie; Niggemann, Monika; Glover, John R.
  • Journal of Biological Chemistry, Vol. 283, Issue 44
  • DOI: 10.1074/jbc.M804849200

Spiral architecture of the Hsp104 disaggregase reveals the basis for polypeptide translocation
journal, August 2016

  • Yokom, Adam L.; Gates, Stephanie N.; Jackrel, Meredith E.
  • Nature Structural & Molecular Biology, Vol. 23, Issue 9
  • DOI: 10.1038/nsmb.3277

Coot model-building tools for molecular graphics
journal, November 2004

  • Emsley, Paul; Cowtan, Kevin
  • Acta Crystallographica Section D Biological Crystallography, Vol. 60, Issue 12, p. 2126-2132
  • DOI: 10.1107/S0907444904019158

Saccharomyces cerevisiae Hsp104 Enhances the Chaperone Capacity of Human Cells and Inhibits Heat Stress-Induced Proapoptotic Signaling
journal, June 2004

  • Mosser, Dick D.; Ho, Sylvia; Glover, John R.
  • Biochemistry, Vol. 43, Issue 25
  • DOI: 10.1021/bi0493766

The Hsp104 N-Terminal Domain Enables Disaggregase Plasticity and Potentiation
journal, March 2015


Conformational Properties of Aggregated Polypeptides Determine ClpB-dependence in the Disaggregation Process
journal, August 2007

  • Lewandowska, Agnieszka; Matuszewska, Marlena; Liberek, Krzysztof
  • Journal of Molecular Biology, Vol. 371, Issue 3
  • DOI: 10.1016/j.jmb.2007.05.057

CryoEM structure of Hsp104 and its mechanistic implication for protein disaggregation
journal, April 2010

  • Lee, S.; Sielaff, B.; Lee, J.
  • Proceedings of the National Academy of Sciences, Vol. 107, Issue 18
  • DOI: 10.1073/pnas.1003572107

PHENIX: a comprehensive Python-based system for macromolecular structure solution
journal, January 2010

  • Adams, Paul D.; Afonine, Pavel V.; Bunkóczi, Gábor
  • Acta Crystallographica Section D Biological Crystallography, Vol. 66, Issue 2, p. 213-221
  • DOI: 10.1107/S0907444909052925

Crystal Structures of the HslVU Peptidase–ATPase Complex Reveal an ATP-Dependent Proteolysis Mechanism
journal, February 2001


Cooperation of Hsp70 and Hsp100 chaperone machines in protein disaggregation
journal, May 2015


Crystal structures of Hsp104 N-terminal domains from Saccharomyces cerevisiae and Candida albicans suggest the mechanism for the function of Hsp104 in dissolving prions
journal, March 2017

  • Wang, Peng; Li, Jingzhi; Weaver, Clarissa
  • Acta Crystallographica Section D Structural Biology, Vol. 73, Issue 4
  • DOI: 10.1107/S2059798317002662

Structural pathway of regulated substrate transfer and threading through an Hsp100 disaggregase
journal, August 2017

  • Deville, Célia; Carroni, Marta; Franke, Kamila B.
  • Science Advances, Vol. 3, Issue 8
  • DOI: 10.1126/sciadv.1701726

NMRPipe: A multidimensional spectral processing system based on UNIX pipes
journal, November 1995

  • Delaglio, Frank; Grzesiek, Stephan; Vuister, GeertenW.
  • Journal of Biomolecular NMR, Vol. 6, Issue 3
  • DOI: 10.1007/BF00197809

Controlled destruction: AAA+ ATPases in protein degradation from bacteria to eukaryotes
journal, April 2009

  • Striebel, Frank; Kress, Wolfgang; Weber-Ban, Eilika
  • Current Opinion in Structural Biology, Vol. 19, Issue 2
  • DOI: 10.1016/j.sbi.2009.02.006

Evidence for an Unfolding/Threading Mechanism for Protein Disaggregation by Saccharomyces cerevisiae Hsp104
journal, May 2004

  • Lum, Ronnie; Tkach, Johnny M.; Vierling, Elizabeth
  • Journal of Biological Chemistry, Vol. 279, Issue 28
  • DOI: 10.1074/jbc.M403777200

MOLREP an Automated Program for Molecular Replacement
journal, December 1997


Structure and mechanism of the hexameric MecA–ClpC molecular machine
journal, March 2011


Structure of the cross-β spine of amyloid-like fibrils
journal, June 2005

  • Nelson, Rebecca; Sawaya, Michael R.; Balbirnie, Melinda
  • Nature, Vol. 435, Issue 7043
  • DOI: 10.1038/nature03680