Comprehensive Structural Characterization of the Human AAA+ Disaggregase CLPB in the Apo- and Substrate-bound States Reveals a Unique Mode of Action Driven by Oligomerization

Overview

Journal PLoS Biol

Specialty Biology

Date 2023 Feb 6

PMID 36745679

Authors

Damu Wu

Yan Liu

YuHao Dai

Guopeng Wang

Guoliang Lu

Yan Chen

Ningning Li

Jinzhong Lin

Ning Gao

Affiliations

Soon will be listed here.

Abstract

The human AAA+ ATPase CLPB (SKD3) is a protein disaggregase in the mitochondrial intermembrane space (IMS) and functions to promote the solubilization of various mitochondrial proteins. Loss-of-function CLPB mutations are associated with a few human diseases with neutropenia and neurological disorders. Unlike canonical AAA+ proteins, CLPB contains a unique ankyrin repeat domain (ANK) at its N-terminus. How CLPB functions as a disaggregase and the role of its ANK domain are currently unclear. Herein, we report a comprehensive structural characterization of human CLPB in both the apo- and substrate-bound states. CLPB assembles into homo-tetradecamers in apo-state and is remodeled into homo-dodecamers upon substrate binding. Conserved pore-loops (PLs) on the ATPase domains form a spiral staircase to grip and translocate the substrate in a step-size of 2 amino acid residues. The ANK domain is not only responsible for maintaining the higher-order assembly but also essential for the disaggregase activity. Interactome analysis suggests that the ANK domain may directly interact with a variety of mitochondrial substrates. These results reveal unique properties of CLPB as a general disaggregase in mitochondria and highlight its potential as a target for the treatment of various mitochondria-related diseases.

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References

Santosh V, Musayev F, Jaiswal R, Zarate-Perez F, Vandewinkel B, Dierckx C . The Cryo-EM structure of AAV2 Rep68 in complex with ssDNA reveals a malleable AAA+ machine that can switch between oligomeric states. Nucleic Acids Res. 2020; 48(22):12983-12999. PMC: 7736791. DOI: 10.1093/nar/gkaa1133. View

Huttlin E, Bruckner R, Navarrete-Perea J, Cannon J, Baltier K, Gebreab F . Dual proteome-scale networks reveal cell-specific remodeling of the human interactome. Cell. 2021; 184(11):3022-3040.e28. PMC: 8165030. DOI: 10.1016/j.cell.2021.04.011. View

Chen X, Glytsou C, Zhou H, Narang S, Reyna D, Lopez A . Targeting Mitochondrial Structure Sensitizes Acute Myeloid Leukemia to Venetoclax Treatment. Cancer Discov. 2019; 9(7):890-909. PMC: 6606342. DOI: 10.1158/2159-8290.CD-19-0117. View

Puchades C, Sandate C, Lander G . The molecular principles governing the activity and functional diversity of AAA+ proteins. Nat Rev Mol Cell Biol. 2019; 21(1):43-58. PMC: 9402527. DOI: 10.1038/s41580-019-0183-6. View

Yu X, VanLoock M, Poplawski A, Kelman Z, Xiang T, Tye B . The Methanobacterium thermoautotrophicum MCM protein can form heptameric rings. EMBO Rep. 2002; 3(8):792-7. PMC: 1084214. DOI: 10.1093/embo-reports/kvf160. View

Cooney I, Han H, Stewart M, Carson R, Hansen D, Iwasa J . Structure of the Cdc48 segregase in the act of unfolding an authentic substrate. Science. 2019; 365(6452):502-505. PMC: 7362759. DOI: 10.1126/science.aax0486. View

Adams P, Afonine P, Bunkoczi G, Chen V, Davis I, Echols N . PHENIX: a comprehensive Python-based system for macromolecular structure solution. Acta Crystallogr D Biol Crystallogr. 2010; 66(Pt 2):213-21. PMC: 2815670. DOI: 10.1107/S0907444909052925. View

Frezza C, Cipolat S, Scorrano L . Organelle isolation: functional mitochondria from mouse liver, muscle and cultured fibroblasts. Nat Protoc. 2007; 2(2):287-95. DOI: 10.1038/nprot.2006.478. View

Kiykim A, Garncarz W, Karakoc-Aydiner E, Ozen A, Kiykim E, Yesil G . Novel CLPB mutation in a patient with 3-methylglutaconic aciduria causing severe neurological involvement and congenital neutropenia. Clin Immunol. 2016; 165:1-3. DOI: 10.1016/j.clim.2016.02.008. View

10.

Kirstein J, Moliere N, Dougan D, Turgay K . Adapting the machine: adaptor proteins for Hsp100/Clp and AAA+ proteases. Nat Rev Microbiol. 2009; 7(8):589-99. DOI: 10.1038/nrmicro2185. View

11.

Pudova E, Krasnov G, Kobelyatskaya A, Savvateeva M, Fedorova M, Pavlov V . Gene Expression Changes and Associated Pathways Involved in the Progression of Prostate Cancer Advanced Stages. Front Genet. 2021; 11:613162. PMC: 7859645. DOI: 10.3389/fgene.2020.613162. View

12.

Santo-Domingo J, Demaurex N . Perspectives on: SGP symposium on mitochondrial physiology and medicine: the renaissance of mitochondrial pH. J Gen Physiol. 2012; 139(6):415-23. PMC: 3362525. DOI: 10.1085/jgp.201110767. View

13.

Fei X, Bell T, Jenni S, Stinson B, Baker T, Harrison S . Structures of the ATP-fueled ClpXP proteolytic machine bound to protein substrate. Elife. 2020; 9. PMC: 7112951. DOI: 10.7554/eLife.52774. View

14.

Cupo R, Rizo A, Braun G, Tse E, Chuang E, Gupta K . Unique structural features govern the activity of a human mitochondrial AAA+ disaggregase, Skd3. Cell Rep. 2022; 40(13):111408. PMC: 9584538. DOI: 10.1016/j.celrep.2022.111408. View

15.

Warren J, Cupo R, Wattanasirakul P, Spencer D, Locke A, Makaryan V . Heterozygous variants of CLPB are a cause of severe congenital neutropenia. Blood. 2021; 139(5):779-791. PMC: 8814677. DOI: 10.1182/blood.2021010762. View

16.

Evans P . An introduction to data reduction: space-group determination, scaling and intensity statistics. Acta Crystallogr D Biol Crystallogr. 2011; 67(Pt 4):282-92. PMC: 3069743. DOI: 10.1107/S090744491003982X. View

17.

Pettersen E, Goddard T, Huang C, Couch G, Greenblatt D, Meng E . UCSF Chimera--a visualization system for exploratory research and analysis. J Comput Chem. 2004; 25(13):1605-12. DOI: 10.1002/jcc.20084. View

18.

Kanabus M, Shahni R, Saldanha J, Murphy E, Plagnol V, Vant Hoff W . Bi-allelic CLPB mutations cause cataract, renal cysts, nephrocalcinosis and 3-methylglutaconic aciduria, a novel disorder of mitochondrial protein disaggregation. J Inherit Metab Dis. 2015; 38(2):211-9. DOI: 10.1007/s10545-015-9813-0. View

19.

Zhang K . Gctf: Real-time CTF determination and correction. J Struct Biol. 2015; 193(1):1-12. PMC: 4711343. DOI: 10.1016/j.jsb.2015.11.003. View

20.

Cupo R, Shorter J . Skd3 (human ClpB) is a potent mitochondrial protein disaggregase that is inactivated by 3-methylglutaconic aciduria-linked mutations. Elife. 2020; 9. PMC: 7343390. DOI: 10.7554/eLife.55279. View