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- EMDB-61847: CryoEM structure of M. tuberculosis ClpP1P2 bound to bortezomib -

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Basic information

Entry
Database: EMDB / ID: EMD-61847
TitleCryoEM structure of M. tuberculosis ClpP1P2 bound to bortezomib
Map data
Sample
  • Complex: CryoEM structure of M. tuberculosis BTZ-ClpP1P2 complex
    • Protein or peptide: ATP-dependent Clp protease proteolytic subunit 1
    • Protein or peptide: ATP-dependent Clp protease proteolytic subunit 2
  • Ligand: N-[(1R)-1-(DIHYDROXYBORYL)-3-METHYLBUTYL]-N-(PYRAZIN-2-YLCARBONYL)-L-PHENYLALANINAMIDE
KeywordsMycobacterium tuberculosis / Caseinolytic protease system / Activation / HYDROLASE
Function / homology
Function and homology information


endopeptidase Clp / endopeptidase Clp complex / ATP-dependent peptidase activity / protein quality control for misfolded or incompletely synthesized proteins / 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 / ClpP/crotonase-like domain superfamily
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) / Mycobacterium tuberculosis H37Rv (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.56 Å
AuthorsZhou B / Zhao H / Gao Y / Chen W / Zhang T / He J / Xiong X
Funding support China, 9 items
OrganizationGrant numberCountry
Other government2021YFA1300903
Other government2021YFA1300904
National Natural Science Foundation of China (NSFC)32300152 China
National Natural Science Foundation of China (NSFC)81973372 China
National Natural Science Foundation of China (NSFC)32170189 China
National Natural Science Foundation of China (NSFC)32241021 China
National Natural Science Foundation of China (NSFC)82341085 China
Other government2022A1515110505
Other government2022M723164
CitationJournal: Nat Commun / Year: 2025
Title: Structural Insights into Bortezomib-Induced Activation of the Caseinolytic Chaperone-Protease System in Mycobacterium tuberculosis.
Authors: Biao Zhou / Yamin Gao / Heyu Zhao / Banghui Liu / Han Zhang / Cuiting Fang / Hang Yuan / Jingjing Wang / Zimu Li / Yi Zhao / Xiaodong Huang / Xiyue Wang / A Sofia F Oliveira / James Spencer ...Authors: Biao Zhou / Yamin Gao / Heyu Zhao / Banghui Liu / Han Zhang / Cuiting Fang / Hang Yuan / Jingjing Wang / Zimu Li / Yi Zhao / Xiaodong Huang / Xiyue Wang / A Sofia F Oliveira / James Spencer / Adrian J Mulholland / Steven G Burston / Jinxing Hu / Ning Su / Xinwen Chen / Jun He / Tianyu Zhang / Xiaoli Xiong /
Abstract: The caseinolytic protease (Clp) system has recently emerged as a promising anti-tuberculosis target. The anti-cancer drug bortezomib exhibits potent anti-mycobacterial activity and binds to ...The caseinolytic protease (Clp) system has recently emerged as a promising anti-tuberculosis target. The anti-cancer drug bortezomib exhibits potent anti-mycobacterial activity and binds to Mycobacterium tuberculosis (Mtb) Clp protease complexes. We determine cryo-EM structures of Mtb ClpP1P2, ClpC1P1P2 and ClpXP1P2 complexes bound to bortezomib in different conformations. Structural and biochemical data indicate that sub-stoichiometric binding by bortezomib to the protease active sites orthosterically activates the MtbClpP1P2 complex. Bortezomib activation of MtbClpP1P2 induces structural changes promoting the recruitment of the chaperone-unfoldases, MtbClpC1 or MtbClpX, facilitating holoenzyme formation. The structures of the MtbClpC1P1P2 holoenzyme indicate that MtbClpC1 motion, induced by ATP rebinding at the MtbClpC1 spiral seam, translocates the substrate. In the MtbClpXP1P2 holoenzyme structure, we identify a specialized substrate channel gating mechanism involving the MtbClpX pore-2 loop and MtbClpP2 N-terminal domains. Our results provide insights into the intricate regulation of the Mtb Clp system and suggest that bortezomib can disrupt this regulation by sub-stoichiometric binding at the Mtb Clp protease sites.
History
DepositionOct 9, 2024-
Header (metadata) releaseMar 19, 2025-
Map releaseMar 19, 2025-
UpdateApr 8, 2026-
Current statusApr 8, 2026Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_61847.png

Downloads & links

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Map

FileDownload / File: emd_61847.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
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AxesZ (Sec.)Y (Row.)X (Col.)
0.71 Å/pix.
x 256 pix.
= 181.76 Å		0.71 Å/pix.
x 256 pix.
= 181.76 Å		0.71 Å/pix.
x 256 pix.
= 181.76 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.71 Å
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Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.06
Minimum - Maximum-0.1123101 - 0.26274982
Average (Standard dev.)0.0014311482 (±0.017233713)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 181.76 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_61847_msk_1.map
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Half map: #2

Fileemd_61847_half_map_1.map
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Half map: #1

Fileemd_61847_half_map_2.map
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Sample components

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Entire : CryoEM structure of M. tuberculosis BTZ-ClpP1P2 complex

EntireName: CryoEM structure of M. tuberculosis BTZ-ClpP1P2 complex
Components
  • Complex: CryoEM structure of M. tuberculosis BTZ-ClpP1P2 complex
    • Protein or peptide: ATP-dependent Clp protease proteolytic subunit 1
    • Protein or peptide: ATP-dependent Clp protease proteolytic subunit 2
  • Ligand: N-[(1R)-1-(DIHYDROXYBORYL)-3-METHYLBUTYL]-N-(PYRAZIN-2-YLCARBONYL)-L-PHENYLALANINAMIDE

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Supramolecule #1: CryoEM structure of M. tuberculosis BTZ-ClpP1P2 complex

SupramoleculeName: CryoEM structure of M. tuberculosis BTZ-ClpP1P2 complex
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Source (natural)Organism: Mycobacterium tuberculosis (bacteria)

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Macromolecule #1: ATP-dependent Clp protease proteolytic subunit 1

MacromoleculeName: ATP-dependent Clp protease proteolytic subunit 1 / type: protein_or_peptide / ID: 1 / Number of copies: 7 / Enantiomer: LEVO / EC number: endopeptidase Clp
Source (natural)Organism: Mycobacterium tuberculosis H37Rv (bacteria) / Strain: H37Rv
Molecular weightTheoretical: 19.383111 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString:
SLTDSVYERL LSERIIFLGS EVNDEIANRL CAQILLLAAE DASKDISLYI NSPGGSISAG MAIYDTMVLA PCDIATYAMG MAASMGEFL LAAGTKGKRY ALPHARILMH QPLGGVTGSA ADIAIQAEQF AVIKKEMFRL NAEFTGQPIE RIEADSDRDR W FTAAEALE YGFVDHIITR

UniProtKB: ATP-dependent Clp protease proteolytic subunit 1

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Macromolecule #2: ATP-dependent Clp protease proteolytic subunit 2

MacromoleculeName: ATP-dependent Clp protease proteolytic subunit 2 / type: protein_or_peptide / ID: 2 / Number of copies: 7 / Enantiomer: LEVO / EC number: endopeptidase Clp
Source (natural)Organism: Mycobacterium tuberculosis H37Rv (bacteria) / Strain: H37Rv
Molecular weightTheoretical: 21.43041 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: LPSFIEHSSF GVKESNPYNK LFEERIIFLG VQVDDASAND IMAQLLVLES LDPDRDITMY INSPGGGFTS LMAIYDTMQY VRADIQTVC LGQAASAAAV LLAAGTPGKR MALPNARVLI HQPSLSGVIQ GQFSDLEIQA AEIERMRTLM ETTLARHTGK D AGVIRKDT ...String:
LPSFIEHSSF GVKESNPYNK LFEERIIFLG VQVDDASAND IMAQLLVLES LDPDRDITMY INSPGGGFTS LMAIYDTMQY VRADIQTVC LGQAASAAAV LLAAGTPGKR MALPNARVLI HQPSLSGVIQ GQFSDLEIQA AEIERMRTLM ETTLARHTGK D AGVIRKDT DRDKILTAEE AKDYGIIDTV LEYRKLS

UniProtKB: ATP-dependent Clp protease proteolytic subunit 2

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Macromolecule #3: N-[(1R)-1-(DIHYDROXYBORYL)-3-METHYLBUTYL]-N-(PYRAZIN-2-YLCARBONYL...

MacromoleculeName: N-[(1R)-1-(DIHYDROXYBORYL)-3-METHYLBUTYL]-N-(PYRAZIN-2-YLCARBONYL)-L-PHENYLALANINAMIDE
type: ligand / ID: 3 / Number of copies: 14 / Formula: BO2
Molecular weightTheoretical: 384.237 Da
Chemical component information

ChemComp-BO2:
N-[(1R)-1-(DIHYDROXYBORYL)-3-METHYLBUTYL]-N-(PYRAZIN-2-YLCARBONYL)-L-PHENYLALANINAMIDE / medication*YM

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 7.5
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.4 µm / Nominal defocus min: 0.8 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

6vgq

Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.56 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 668234
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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