+Open data
-Basic information
Entry | Database: PDB / ID: 6lsy | |||||||||||||||
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Title | AAA+ ATPase, ClpL from Streptococcus pneumoniae - ATP bound | |||||||||||||||
Components | ATP-dependent Clp protease, ATP-binding subunit | |||||||||||||||
Keywords | CHAPERONE / AAA+ ATPase / Streptococcus pneumoniae | |||||||||||||||
Function / homology | Function and homology information | |||||||||||||||
Biological species | Streptococcus pneumoniae (bacteria) | |||||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 6.33 Å | |||||||||||||||
Authors | Kim, G. / Lee, S.G. / Han, S. / Jung, J. / Jeong, H.S. / Hyun, J.K. / Rhee, D.K. / Kim, H.M. / Lee, S. | |||||||||||||||
Funding support | Korea, Republic Of, 4items
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Citation | Journal: FASEB J / Year: 2020 Title: ClpL is a functionally active tetradecameric AAA+ chaperone, distinct from hexameric/dodecameric ones. Authors: Gyuhee Kim / Seong-Gyu Lee / Seungsu Han / Jaeeun Jung / Hyeong Seop Jeong / Jae-Kyung Hyun / Dong-Kwon Rhee / Ho Min Kim / Sangho Lee / Abstract: AAA+ (ATPases associated with diverse cellular activities) chaperones are involved in a plethora of cellular activities to ensure protein homeostasis. The function of AAA+ chaperones is mostly ...AAA+ (ATPases associated with diverse cellular activities) chaperones are involved in a plethora of cellular activities to ensure protein homeostasis. The function of AAA+ chaperones is mostly modulated by their hexameric/dodecameric quaternary structures. Here we report the structural and biochemical characterizations of a tetradecameric AAA+ chaperone, ClpL from Streptococcus pneumoniae. ClpL exists as a tetradecamer in solution in the presence of ATP. The cryo-EM structure of ClpL at 4.5 Å resolution reveals a striking tetradecameric arrangement. Solution structures of ClpL derived from small-angle X-ray scattering data suggest that the tetradecameric ClpL could assume a spiral conformation found in active hexameric/dodecameric AAA+ chaperone structures. Vertical positioning of the middle domain accounts for the head-to-head arrangement of two heptameric rings. Biochemical activity assays with site-directed mutagenesis confirmed the critical roles of residues both in the integrity of the tetradecameric arrangement and activities of ClpL. Non-conserved Q321 and R670 are crucial in the heptameric ring assembly of ClpL. These results establish that ClpL is a functionally active tetradecamer, clearly distinct from hexameric/dodecameric AAA+ chaperones. | |||||||||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 6lsy.cif.gz | 2.7 MB | Display | PDBx/mmCIF format |
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PDB format | pdb6lsy.ent.gz | Display | PDB format | |
PDBx/mmJSON format | 6lsy.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/ls/6lsy ftp://data.pdbj.org/pub/pdb/validation_reports/ls/6lsy | HTTPS FTP |
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-Related structure data
Related structure data | 0965MC 0967C 6lt4C M: map data used to model this data C: citing same article (ref.) |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
#1: Protein | Mass: 77599.094 Da / Num. of mol.: 14 / Mutation: E193A/E526A Source method: isolated from a genetically manipulated source Source: (gene. exp.) Streptococcus pneumoniae (bacteria) / Gene: clpL, SPRM200_0307 / Production host: Escherichia coli (E. coli) / References: UniProt: A0A2U3RY34 |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: ClpL Trap(E193A/E526A):ATP-bound / Type: ORGANELLE OR CELLULAR COMPONENT / Entity ID: all / Source: RECOMBINANT | |||||||||||||||||||||||||
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Molecular weight | Value: 1.1 MDa / Experimental value: YES | |||||||||||||||||||||||||
Source (natural) | Organism: Streptococcus pneumoniae (bacteria) | |||||||||||||||||||||||||
Source (recombinant) | Organism: Escherichia coli (E. coli) | |||||||||||||||||||||||||
Buffer solution | pH: 7.5 | |||||||||||||||||||||||||
Buffer component |
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Specimen | Conc.: 0.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | |||||||||||||||||||||||||
Specimen support | Grid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3 | |||||||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: DARK FIELD |
Image recording | Electron dose: 60 e/Å2 / Film or detector model: FEI FALCON III (4k x 4k) |
-Processing
Software |
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EM software |
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CTF correction | Type: NONE | ||||||||||||||||||||||||
Particle selection | Num. of particles selected: 104148 | ||||||||||||||||||||||||
Symmetry | Point symmetry: D7 (2x7 fold dihedral) | ||||||||||||||||||||||||
3D reconstruction | Resolution: 6.33 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 52689 / Symmetry type: POINT | ||||||||||||||||||||||||
Atomic model building | Protocol: AB INITIO MODEL | ||||||||||||||||||||||||
Refinement | Cross valid method: NONE / Stereochemistry target values: GeoStd + Monomer Library | ||||||||||||||||||||||||
Refine LS restraints |
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