The cytoplasmic domain of the AAA+ protease FtsH is tilted with respect to the membrane to facilitate substrate entry.
ATP-dependent protease
conformational change
electron microscopy
membrane protein
protein structure
Journal
The Journal of biological chemistry
ISSN: 1083-351X
Titre abrégé: J Biol Chem
Pays: United States
ID NLM: 2985121R
Informations de publication
Date de publication:
Historique:
received:
25
06
2020
revised:
28
10
2020
accepted:
05
11
2020
pubmed:
7
11
2020
medline:
28
8
2021
entrez:
6
11
2020
Statut:
ppublish
Résumé
AAA+ proteases are degradation machines that use ATP hydrolysis to unfold protein substrates and translocate them through a central pore toward a degradation chamber. FtsH, a bacterial membrane-anchored AAA+ protease, plays a vital role in membrane protein quality control. How substrates reach the FtsH central pore is an open key question that is not resolved by the available atomic structures of cytoplasmic and periplasmic domains. In this work, we used both negative stain TEM and cryo-EM to determine 3D maps of the full-length Aquifex aeolicus FtsH protease. Unexpectedly, we observed that detergent solubilization induces the formation of fully active FtsH dodecamers, which consist of two FtsH hexamers in a single detergent micelle. The striking tilted conformation of the cytosolic domain in the FtsH dodecamer visualized by negative stain TEM suggests a lateral substrate entrance between the membrane and cytosolic domain. Such a substrate path was then resolved in the cryo-EM structure of the FtsH hexamer. By mapping the available structural information and structure predictions for the transmembrane helices to the amino acid sequence we identified a linker of ∼20 residues between the second transmembrane helix and the cytosolic domain. This unique polypeptide appears to be highly flexible and turned out to be essential for proper functioning of FtsH as its deletion fully eliminated the proteolytic activity of FtsH.
Identifiants
pubmed: 33154162
pii: S0021-9258(20)00015-0
doi: 10.1074/jbc.RA120.014739
pmc: PMC7949044
pii:
doi:
Substances chimiques
Metalloendopeptidases
EC 3.4.24.-
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
100029Informations de copyright
Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.
Déclaration de conflit d'intérêts
Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article.
Références
FEBS Lett. 2012 Sep 21;586(19):3117-21
pubmed: 23166924
Sci Rep. 2016 Sep 16;6:33631
pubmed: 27632940
J Struct Biol. 2007 Jan;157(1):38-46
pubmed: 16859925
Structure. 2002 Oct;10(10):1415-23
pubmed: 12377127
Bioinformatics. 2001 Sep;17(9):849-50
pubmed: 11590105
J Struct Biol. 2016 Jul;195(1):93-9
pubmed: 27108186
Nucleic Acids Res. 1997 Sep 1;25(17):3389-402
pubmed: 9254694
Elife. 2016 Dec 21;5:
pubmed: 28001128
Nucleic Acids Res. 2016 Jul 8;44(W1):W344-50
pubmed: 27166375
Mol Cell Biol. 2006 Nov;26(22):8488-97
pubmed: 16966379
J Comput Chem. 2004 Oct;25(13):1605-12
pubmed: 15264254
Methods. 2008 Oct;46(2):73-82
pubmed: 18625320
J Struct Biol. 2015 Nov;192(2):216-21
pubmed: 26278980
Biochemistry. 2016 Oct 11;55(40):5649-5652
pubmed: 27677373
Proc Natl Acad Sci U S A. 2009 Dec 22;106(51):21579-84
pubmed: 19955424
Biochim Biophys Acta Biomembr. 2019 May 1;1861(5):939-957
pubmed: 30776334
J Biol Chem. 2011 Feb 11;286(6):4404-11
pubmed: 21147776
EMBO J. 1995 Jun 1;14(11):2551-60
pubmed: 7781608
J Synchrotron Radiat. 2008 May;15(Pt 3):208-10
pubmed: 18421140
Curr Res Struct Biol. 2019 Oct 23;1:13-20
pubmed: 34235464
J Mol Biol. 2015 Feb 27;427(4):910-923
pubmed: 25576874
Nucleic Acids Res. 2016 Jul 8;44(W1):W449-54
pubmed: 27131374
J Struct Biol. 2007 Jan;157(1):117-25
pubmed: 16828314
J Struct Biol. 2017 May;198(2):124-133
pubmed: 28344036
Mol Cell. 2006 Jun 9;22(5):575-85
pubmed: 16762831
J Bacteriol. 2013 May;195(9):1912-9
pubmed: 23417489
J Bacteriol. 2011 Sep;193(18):4790-7
pubmed: 21764915
J Biol Chem. 2013 Feb 15;288(7):4792-8
pubmed: 23283966
Nucleic Acids Res. 2015 Jul 1;43(W1):W408-12
pubmed: 25943549
Trends Biotechnol. 1998 Aug;16(8):329-32
pubmed: 9720321
Biochemistry. 2003 Sep 16;42(36):10843-52
pubmed: 12962509
Nucleic Acids Res. 2005 Jul 1;33(Web Server issue):W72-6
pubmed: 15980571
Proc Natl Acad Sci U S A. 2006 Feb 28;103(9):3066-71
pubmed: 16484367
Sci Adv. 2020 May 20;6(21):eaba8404
pubmed: 32490208
J Biol Chem. 2008 Nov 7;283(45):30433-7
pubmed: 18650443
J Med Genet. 2005 Jul;42(7):529-39
pubmed: 15994873
J Struct Biol. 2005 Oct;152(1):36-51
pubmed: 16182563
Nat Methods. 2010 Dec;7(12):1003-8
pubmed: 21037590
Acta Crystallogr D Biol Crystallogr. 2015 Jun;71(Pt 6):1307-18
pubmed: 26057670
Proc Natl Acad Sci U S A. 2002 Jun 11;99(12):8066-71
pubmed: 12034886
Elife. 2018 Nov 09;7:
pubmed: 30412051
Biol Chem. 2017 May 1;398(5-6):625-635
pubmed: 28085670
J Mol Biol. 2001 Jan 19;305(3):567-80
pubmed: 11152613
Mitochondrion. 2019 Nov;49:121-127
pubmed: 31377246
Science. 2017 Nov 3;358(6363):
pubmed: 29097521
Sci Rep. 2017 Feb 08;7:41751
pubmed: 28176812
Nat Rev Microbiol. 2016 Jan;14(1):33-44
pubmed: 26639779
FEBS Open Bio. 2019 Sep;9(9):1536-1551
pubmed: 31237118
Cell. 2005 Oct 21;123(2):277-89
pubmed: 16239145
Nat Methods. 2017 Apr;14(4):331-332
pubmed: 28250466
Nucleic Acids Res. 2009 Jul;37(Web Server issue):W575-80
pubmed: 19465378
Proc Natl Acad Sci U S A. 2013 Apr 30;110(18):7264-9
pubmed: 23589842
Nat Methods. 2009 May;6(5):343-5
pubmed: 19363495
Nucleic Acids Res. 2004 Jul 1;32(Web Server issue):W321-6
pubmed: 15215403
J Struct Biol. 1996 Jan-Feb;116(1):71-6
pubmed: 8742726
FEBS Lett. 2010 Jan 21;584(2):252-64
pubmed: 19931533
Methods Enzymol. 2016;579:159-89
pubmed: 27572727
Nucleic Acids Res. 2015 Jul 1;43(W1):W401-7
pubmed: 25969446
Nature. 2015 Feb 5;518(7537):61-7
pubmed: 25581794
J Struct Biol. 2012 Dec;180(3):519-30
pubmed: 23000701