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- EMDB-38537: Cryo-EM structure of ClpP1P2 in complex with ADEP1 from Streptomy... -

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Basic information

Entry
Database: EMDB / ID: EMD-38537
TitleCryo-EM structure of ClpP1P2 in complex with ADEP1 from Streptomyces hawaiiensis
Map data
Sample
  • Complex: ClpP1,ClpP2
    • Protein or peptide: ATP-dependent Clp protease proteolytic subunit
    • Protein or peptide: ATP-dependent Clp protease proteolytic subunit
    • Protein or peptide: ADEP1
Keywordsprotease / protein degradation / proteostasis / proteolysis / HYDROLASE
Function / homology
Function and homology information


endopeptidase Clp / ATP-dependent peptidase activity / serine-type endopeptidase activity / proteolysis / cytoplasm
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 / ATP-dependent Clp protease proteolytic subunit
Similarity search - Component
Biological speciesStreptomyces hawaiiensis (bacteria) / synthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.8 Å
AuthorsXu X / Long F
Funding support China, 1 items
OrganizationGrant numberCountry
Ministry of Science and Technology (MoST, China)2021YFA0909500 China
CitationJournal: mBio / Year: 2024
Title: Structural insights into the Clp protein degradation machinery.
Authors: Xiaolong Xu / Yanhui Wang / Wei Huang / Danyang Li / Zixin Deng / Feng Long /
Abstract: The Clp protease system is important for maintaining proteostasis in bacteria. It consists of ClpP serine proteases and an AAA+ Clp-ATPase such as ClpC1. The hexameric ATPase ClpC1 utilizes the ...The Clp protease system is important for maintaining proteostasis in bacteria. It consists of ClpP serine proteases and an AAA+ Clp-ATPase such as ClpC1. The hexameric ATPase ClpC1 utilizes the energy of ATP binding and hydrolysis to engage, unfold, and translocate substrates into the proteolytic chamber of homo- or hetero-tetradecameric ClpP for degradation. The assembly between the hetero-tetradecameric ClpP1P2 chamber and the Clp-ATPases containing tandem ATPase domains from the same species has not been studied in depth. Here, we present cryo-EM structures of the substrate-bound ClpC1:shClpP1P2 from , and shClpP1P2 in complex with ADEP1, a natural compound produced by and known to cause over-activation and dysregulation of the ClpP proteolytic core chamber. Our structures provide detailed information on the shClpP1-shClpP2, shClpP2-ClpC1, and ADEP1-shClpP1/P2 interactions, reveal conformational transition of ClpC1 during the substrate translocation, and capture a rotational ATP hydrolysis mechanism likely dominated by the D1 ATPase activity of chaperones.IMPORTANCEThe Clp-dependent proteolysis plays an important role in bacterial homeostasis and pathogenesis. The ClpP protease system is an effective drug target for antibacterial therapy. can produce a class of potent acyldepsipeptide antibiotics such as ADEP1, which could affect the ClpP protease activity. Although hosts one of the most intricate ClpP systems in nature, very little was known about its Clp protease mechanism and the impact of ADEP molecules on ClpP. The significance of our research is in dissecting the functional mechanism of the assembled Clp degradation machinery, as well as the interaction between ADEP1 and the ClpP proteolytic chamber, by solving high-resolution structures of the substrate-bound Clp system in . The findings shed light on our understanding of the Clp-dependent proteolysis in bacteria, which will enhance the development of antimicrobial drugs targeting the Clp protease system, and help fighting against bacterial multidrug resistance.
History
DepositionJan 2, 2024-
Header (metadata) releaseMar 27, 2024-
Map releaseMar 27, 2024-
UpdateApr 24, 2024-
Current statusApr 24, 2024Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_38537.png

Downloads & links

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Map

FileDownload / File: emd_38537.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 0.84 Å
Density
Contour LevelBy AUTHOR: 0.15
Minimum - Maximum-0.1250543 - 0.3024761
Average (Standard dev.)0.0017654485 (±0.017999994)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 215.04 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_38537_msk_1.map
Projections & Slices
AxesZYX

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Additional map: sharpened

Fileemd_38537_additional_1.map
Annotationsharpened
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Half map: #2

Fileemd_38537_half_map_1.map
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Half map: #1

Fileemd_38537_half_map_2.map
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Sample components

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Entire : ClpP1,ClpP2

EntireName: ClpP1,ClpP2
Components
  • Complex: ClpP1,ClpP2
    • Protein or peptide: ATP-dependent Clp protease proteolytic subunit
    • Protein or peptide: ATP-dependent Clp protease proteolytic subunit
    • Protein or peptide: ADEP1

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Supramolecule #1: ClpP1,ClpP2

SupramoleculeName: ClpP1,ClpP2 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Streptomyces hawaiiensis (bacteria)

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Macromolecule #1: ATP-dependent Clp protease proteolytic subunit

MacromoleculeName: ATP-dependent Clp protease proteolytic subunit / type: protein_or_peptide / ID: 1 / Number of copies: 7 / Enantiomer: LEVO / EC number: endopeptidase Clp
Source (natural)Organism: Streptomyces hawaiiensis (bacteria)
Molecular weightTheoretical: 22.632605 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MGSSHHHHHH SSGLVPRGSH MGLGDQVYNR LLNERIIFLG QPVDDDIANK ITAQLLLLAS DPEKDIYLYI NSPGGSITAG MAIYDTMQY IKNDVVTIAM GLAAAMGQFL LSAGTPGKRF ALPNAEILIH QPSAGLAGSA SDIKIHAERL LHTKKRMAEL T SQHTGQTI ...String:
MGSSHHHHHH SSGLVPRGSH MGLGDQVYNR LLNERIIFLG QPVDDDIANK ITAQLLLLAS DPEKDIYLYI NSPGGSITAG MAIYDTMQY IKNDVVTIAM GLAAAMGQFL LSAGTPGKRF ALPNAEILIH QPSAGLAGSA SDIKIHAERL LHTKKRMAEL T SQHTGQTI EQITRDSDRD RWFDAFEAKE YGLIDDVMTT AAGMPGGGGT GA

UniProtKB: ATP-dependent Clp protease proteolytic subunit

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Macromolecule #2: ATP-dependent Clp protease proteolytic subunit

MacromoleculeName: ATP-dependent Clp protease proteolytic subunit / type: protein_or_peptide / ID: 2 / Number of copies: 7 / Enantiomer: LEVO / EC number: endopeptidase Clp
Source (natural)Organism: Streptomyces hawaiiensis (bacteria)
Molecular weightTheoretical: 24.05824 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MGSSHHHHHH SSGLVPRGSH MASMTGGQQM GRGSEYDPYA KLFEERVIFL GVQIDDASAN DVMAQLLCLE SMDPDRDISV YINSPGGSF TALTAIYDTM QYVKPDVQTV CMGQAAAAAA VLLAAGTPGK RMALPNARVL IHQPYSETGR GQVSDLEIAA N EILRMRSQ ...String:
MGSSHHHHHH SSGLVPRGSH MASMTGGQQM GRGSEYDPYA KLFEERVIFL GVQIDDASAN DVMAQLLCLE SMDPDRDISV YINSPGGSF TALTAIYDTM QYVKPDVQTV CMGQAAAAAA VLLAAGTPGK RMALPNARVL IHQPYSETGR GQVSDLEIAA N EILRMRSQ LEDMLAKHST TPVEKIREDI ERDKILTAED ALSYGLIDQV ISTRKMDNSS LR

UniProtKB: ATP-dependent Clp protease proteolytic subunit

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Macromolecule #3: ADEP1

MacromoleculeName: ADEP1 / type: protein_or_peptide / ID: 3 / Number of copies: 14 / Enantiomer: LEVO
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 736.855 Da
SequenceString:
(OTT)FSP(MAA)A(MP8)

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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

Concentration3 mg/mL
BufferpH: 7.5
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 50.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: INSILICO MODEL
Initial angle assignmentType: RANDOM ASSIGNMENT
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final reconstructionApplied symmetry - Point group: C7 (7 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 2.8 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 504306
FSC plot (resolution estimation)

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