[English] 日本語
Yorodumi
- PDB-6vgk: ClpP1P2 complex from M. tuberculosis -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: PDB / ID: 6vgk
TitleClpP1P2 complex from M. tuberculosis
Components
  • ATP-dependent Clp protease proteolytic subunit 1
  • ATP-dependent Clp protease proteolytic subunit 2
KeywordsHYDROLASE / Complex / protease / ClpP / tuberculosis
Function / homology
Function and homology information


endopeptidase Clp / endopeptidase Clp complex / protein quality control for misfolded or incompletely synthesized proteins / ATP-dependent peptidase activity / peptidoglycan-based cell wall / ATPase binding / serine-type endopeptidase activity / plasma membrane / cytosol
Similarity search - Function
ClpP, Ser active site / Endopeptidase Clp serine active site. / ClpP, histidine active site / Endopeptidase Clp histidine active site. / ATP-dependent Clp protease proteolytic subunit / Clp protease proteolytic subunit /Translocation-enhancing protein TepA / Clp protease / 2-enoyl-CoA Hydratase; Chain A, domain 1 / 2-enoyl-CoA Hydratase; Chain A, domain 1 / ClpP/crotonase-like domain superfamily ...ClpP, Ser active site / Endopeptidase Clp serine active site. / ClpP, histidine active site / Endopeptidase Clp histidine active site. / ATP-dependent Clp protease proteolytic subunit / Clp protease proteolytic subunit /Translocation-enhancing protein TepA / Clp protease / 2-enoyl-CoA Hydratase; Chain A, domain 1 / 2-enoyl-CoA Hydratase; Chain A, domain 1 / ClpP/crotonase-like domain superfamily / Alpha-Beta Complex / Alpha Beta
Similarity search - Domain/homology
ATP-dependent Clp protease proteolytic subunit 2 / ATP-dependent Clp protease proteolytic subunit 1
Similarity search - Component
Biological speciesMycobacterium tuberculosis (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.1 Å
AuthorsRipstein, Z.A. / Vahidi, S. / Rubinstein, J.L. / Kay, L.E.
Funding support Canada, 2items
OrganizationGrant numberCountry
Canadian Institutes of Health Research (CIHR)FDN-503573 Canada
Canadian Institutes of Health Research (CIHR)PJT-162186 Canada
CitationJournal: Proc Natl Acad Sci U S A / Year: 2020
Title: An allosteric switch regulates ClpP1P2 protease function as established by cryo-EM and methyl-TROSY NMR.
Authors: Siavash Vahidi / Zev A Ripstein / Jordan B Juravsky / Enrico Rennella / Alfred L Goldberg / Anthony K Mittermaier / John L Rubinstein / Lewis E Kay /
Abstract: The 300-kDa ClpP1P2 protease from collaborates with the AAA+ (ATPases associated with a variety of cellular activities) unfoldases, ClpC1 and ClpX, to degrade substrate proteins. Unlike in other ...The 300-kDa ClpP1P2 protease from collaborates with the AAA+ (ATPases associated with a variety of cellular activities) unfoldases, ClpC1 and ClpX, to degrade substrate proteins. Unlike in other bacteria, all of the components of the Clp system are essential for growth and virulence of mycobacteria, and their inhibitors show promise as antibiotics. MtClpP1P2 is unique in that it contains a pair of distinct ClpP1 and ClpP2 rings and also requires the presence of activator peptides, such as benzoyl-leucyl-leucine (Bz-LL), for function. Understanding the structural basis for this requirement has been elusive but is critical for the rational design and improvement of antituberculosis (anti-TB) therapeutics that target the Clp system. Here, we present a combined biophysical and biochemical study to explore the structure-dynamics-function relationship in MtClpP1P2. Electron cryomicroscopy (cryo-EM) structures of apo and acyldepsipeptide-bound MtClpP1P2 explain their lack of activity by showing loss of a key β-sheet in a sequence known as the handle region that is critical for the proper formation of the catalytic triad. Methyl transverse relaxation-optimized spectroscopy (TROSY)-based NMR, cryo-EM, and biochemical assays show that, on binding Bz-LL or covalent inhibitors, MtClpP1P2 undergoes a conformational change from an inactive compact state to an active extended structure that can be explained by a modified Monod-Wyman-Changeux model. Our study establishes a critical role for the handle region as an on/off switch for function and shows extensive allosteric interactions involving both intra- and interring communication that regulate MtClpP1P2 activity and that can potentially be exploited by small molecules to target .
History
DepositionJan 8, 2020Deposition site: RCSB / Processing site: RCSB
Revision 1.0Mar 18, 2020Provider: repository / Type: Initial release
Revision 1.1Apr 1, 2020Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation_author.identifier_ORCID
Revision 1.2Mar 6, 2024Group: Data collection / Database references / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / em_3d_fitting_list / pdbx_initial_refinement_model
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id / _em_3d_fitting_list.source_name / _em_3d_fitting_list.type

-
Structure visualization

Movie
  • Deposited structure unit
  • Imaged by Jmol
  • Download
  • Superimposition on EM map
  • EMDB-21197
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

-
Assembly

Deposited unit
A: ATP-dependent Clp protease proteolytic subunit 2
B: ATP-dependent Clp protease proteolytic subunit 2
C: ATP-dependent Clp protease proteolytic subunit 2
D: ATP-dependent Clp protease proteolytic subunit 2
E: ATP-dependent Clp protease proteolytic subunit 2
F: ATP-dependent Clp protease proteolytic subunit 2
G: ATP-dependent Clp protease proteolytic subunit 2
H: ATP-dependent Clp protease proteolytic subunit 1
I: ATP-dependent Clp protease proteolytic subunit 1
J: ATP-dependent Clp protease proteolytic subunit 1
K: ATP-dependent Clp protease proteolytic subunit 1
L: ATP-dependent Clp protease proteolytic subunit 1
M: ATP-dependent Clp protease proteolytic subunit 1
N: ATP-dependent Clp protease proteolytic subunit 1


Theoretical massNumber of molelcules
Total (without water)300,68414
Polymers300,68414
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: Methyl-Trosy NMR experiments confirmed formation of the complex.
TypeNameSymmetry operationNumber
identity operation1_5551

-
Components

#1: Protein
ATP-dependent Clp protease proteolytic subunit 2 / Endopeptidase Clp 2


Mass: 21888.877 Da / Num. of mol.: 7
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mycobacterium tuberculosis (bacteria) / Gene: clpP2, Rv2460c, MTV008.16c / Plasmid: pET24a+
Details (production host): Added cleavable N-terminal His 6 -SUMO tag
Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P9WPC3, endopeptidase Clp
#2: Protein
ATP-dependent Clp protease proteolytic subunit 1 / Endopeptidase Clp 1


Mass: 21065.934 Da / Num. of mol.: 7
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mycobacterium tuberculosis (bacteria) / Gene: clpP1, clpP, Rv2461c, MTV008.17c / Plasmid: pET24a+
Details (production host): Added cleavable N-terminal His 6 -SUMO tag
Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P9WPC5, endopeptidase Clp

-
Experimental details

-
Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

-
Sample preparation

ComponentName: ClpP1P2 complex / Type: COMPLEX / Details: Complex formed between P1 and P2 heptameric rings / Entity ID: all / Source: RECOMBINANT
Molecular weightValue: 0.3 MDa / Experimental value: NO
Source (natural)Organism: Mycobacterium tuberculosis (bacteria)
Source (recombinant)Organism: Escherichia coli (E. coli) / Strain: Bl21(DE3) / Plasmid: pet24a+
Buffer solutionpH: 7
Details: IGEPAL-CA630 was added shortly prior to vitrification
Buffer component
IDConc.NameBuffer-ID
150 mMImidazole1
2100 mMPotassium Chloride1
35 mMDithiothreitol1
40.025 %IGEPAL-CA6301
SpecimenConc.: 20 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES / Details: Mono-disperse complexes
Specimen supportGrid material: COPPER/RHODIUM / Grid mesh size: 400 divisions/in. / Grid type: Homemade
VitrificationInstrument: FEI VITROBOT MARK III / Cryogen name: ETHANE-PROPANE / Humidity: 100 % / Chamber temperature: 277 K / Details: Blotted for 4.5 seconds at an offset of -5 mm

-
Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal magnification: 75000 X / Nominal defocus max: 2000 nm / Nominal defocus min: 700 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm / Alignment procedure: ZEMLIN TABLEAU
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Temperature (max): 77 K / Temperature (min): 70 K
Image recordingAverage exposure time: 60 sec. / Electron dose: 43 e/Å2 / Detector mode: COUNTING / Film or detector model: FEI FALCON III (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 2092
Image scansWidth: 4096 / Height: 4096

-
Processing

EM software
IDNameVersionCategory
1cryoSPARC2.1particle selection
2EPUimage acquisition
4cryoSPARC2.1CTF correction
7UCSF Chimeramodel fitting
9cryoSPARC2.1initial Euler assignment
10cryoSPARC2.1final Euler assignment
11cryoSPARC2.1classification
12cryoSPARC2.13D reconstruction
13RosettaEMmodel refinement
14PHENIXmodel refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 612408
SymmetryPoint symmetry: C7 (7 fold cyclic)
3D reconstructionResolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 373064 / Algorithm: FOURIER SPACE / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT / Space: REAL
Atomic model buildingPDB-ID: 5DZK

5dzk


Accession code: 5DZK / Source name: PDB / Type: experimental model

+
About Yorodumi

-
News

-
Feb 9, 2022. New format data for meta-information of EMDB entries

New format data for meta-information of EMDB entries

  • Version 3 of the EMDB header file is now the official format.
  • The previous official version 1.9 will be removed from the archive.

Related info.:EMDB header

External links:wwPDB to switch to version 3 of the EMDB data model

-
Aug 12, 2020. Covid-19 info

Covid-19 info

URL: https://pdbj.org/emnavi/covid19.php

New page: Covid-19 featured information page in EM Navigator.

Related info.:Covid-19 info / Mar 5, 2020. Novel coronavirus structure data

+
Mar 5, 2020. Novel coronavirus structure data

Novel coronavirus structure data

Related info.:Yorodumi Speices / Aug 12, 2020. Covid-19 info

External links:COVID-19 featured content - PDBj / Molecule of the Month (242)：Coronavirus Proteases

+
Jan 31, 2019. EMDB accession codes are about to change! (news from PDBe EMDB page)

EMDB accession codes are about to change! (news from PDBe EMDB page)

  • The allocation of 4 digits for EMDB accession codes will soon come to an end. Whilst these codes will remain in use, new EMDB accession codes will include an additional digit and will expand incrementally as the available range of codes is exhausted. The current 4-digit format prefixed with “EMD-” (i.e. EMD-XXXX) will advance to a 5-digit format (i.e. EMD-XXXXX), and so on. It is currently estimated that the 4-digit codes will be depleted around Spring 2019, at which point the 5-digit format will come into force.
  • The EM Navigator/Yorodumi systems omit the EMD- prefix.

Related info.:Q: What is EMD? / ID/Accession-code notation in Yorodumi/EM Navigator

External links:EMDB Accession Codes are Changing Soon! / Contact to PDBj

+
Jul 12, 2017. Major update of PDB

Major update of PDB

  • wwPDB released updated PDB data conforming to the new PDBx/mmCIF dictionary.
  • This is a major update changing the version number from 4 to 5, and with Remediation, in which all the entries are updated.
  • In this update, many items about electron microscopy experimental information are reorganized (e.g. em_software).
  • Now, EM Navigator and Yorodumi are based on the updated data.

External links:wwPDB Remediation / Enriched Model Files Conforming to OneDep Data Standards Now Available in the PDB FTP Archive

-
Yorodumi

Thousand views of thousand structures

  • Yorodumi is a browser for structure data from EMDB, PDB, SASBDB, etc.
  • This page is also the successor to EM Navigator detail page, and also detail information page/front-end page for Omokage search.
  • The word "yorodu" (or yorozu) is an old Japanese word meaning "ten thousand". "mi" (miru) is to see.

Related info.:EMDB / PDB / SASBDB / Comparison of 3 databanks / Yorodumi Search / Aug 31, 2016. New EM Navigator & Yorodumi / Yorodumi Papers / Jmol/JSmol / Function and homology information / Changes in new EM Navigator and Yorodumi

Read more