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Yorodumi- EMDB-4626: ClpB (DWB and K476C mutant) bound to casein in presence of ATPgam... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-4626 | ||||||||||||
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Title | ClpB (DWB and K476C mutant) bound to casein in presence of ATPgammaS - state KC-2A | ||||||||||||
Map data | ClpB (DWB and K476C mutant) bound to a casein substrate in presence of ATPgammaS - State KC2A | ||||||||||||
Sample |
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Keywords | disaggregase / proteostasis / AAA / CHAPERONE | ||||||||||||
Function / homology | Function and homology information cellular response to heat / response to heat / protein refolding / ATP hydrolysis activity / ATP binding / identical protein binding / membrane / cytosol / cytoplasm Similarity search - Function | ||||||||||||
Biological species | Escherichia coli (E. coli) / Bos taurus (cattle) | ||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.8 Å | ||||||||||||
Authors | Deville C / Saibil HR | ||||||||||||
Funding support | United Kingdom, 3 items
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Citation | Journal: Cell Rep / Year: 2019 Title: Two-Step Activation Mechanism of the ClpB Disaggregase for Sequential Substrate Threading by the Main ATPase Motor. Authors: Célia Deville / Kamila Franke / Axel Mogk / Bernd Bukau / Helen R Saibil / Abstract: AAA+ proteins form asymmetric hexameric rings that hydrolyze ATP and thread substrate proteins through a central channel via mobile substrate-binding pore loops. Understanding how ATPase and ...AAA+ proteins form asymmetric hexameric rings that hydrolyze ATP and thread substrate proteins through a central channel via mobile substrate-binding pore loops. Understanding how ATPase and threading activities are regulated and intertwined is key to understanding the AAA+ protein mechanism. We studied the disaggregase ClpB, which contains tandem ATPase domains (AAA1, AAA2) and shifts between low and high ATPase and threading activities. Coiled-coil M-domains repress ClpB activity by encircling the AAA1 ring. Here, we determine the mechanism of ClpB activation by comparing ATPase mechanisms and cryo-EM structures of ClpB wild-type and a constitutively active ClpB M-domain mutant. We show that ClpB activation reduces ATPase cooperativity and induces a sequential mode of ATP hydrolysis in the AAA2 ring, the main ATPase motor. AAA1 and AAA2 rings do not work synchronously but in alternating cycles. This ensures high grip, enabling substrate threading via a processive, rope-climbing mechanism. | ||||||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_4626.map.gz | 85.4 MB | EMDB map data format | |
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Header (meta data) | emd-4626-v30.xml emd-4626.xml | 25.6 KB 25.6 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_4626_fsc.xml | 10.3 KB | Display | FSC data file |
Images | emd_4626.png | 163.7 KB | ||
Filedesc metadata | emd-4626.cif.gz | 7.4 KB | ||
Others | emd_4626_additional.map.gz emd_4626_half_map_1.map.gz emd_4626_half_map_2.map.gz | 57.7 MB 71.3 MB 29.7 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-4626 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-4626 | HTTPS FTP |
-Validation report
Summary document | emd_4626_validation.pdf.gz | 468.4 KB | Display | EMDB validaton report |
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Full document | emd_4626_full_validation.pdf.gz | 467.5 KB | Display | |
Data in XML | emd_4626_validation.xml.gz | 15.5 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-4626 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-4626 | HTTPS FTP |
-Related structure data
Related structure data | 6qs7MC 4621C 4622C 4623C 4624C 4625C 4627C 4940C 4941C 4942C 6qs4C 6qs6C 6qs8C 6rn2C 6rn3C 6rn4C C: citing same article (ref.) M: atomic model generated by this map |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_4626.map.gz / Format: CCP4 / Size: 91.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | ClpB (DWB and K476C mutant) bound to a casein substrate in presence of ATPgammaS - State KC2A | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.05 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Additional map: map filtered according to local resolution
File | emd_4626_additional.map | ||||||||||||
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Annotation | map filtered according to local resolution | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Half map 1
File | emd_4626_half_map_1.map | ||||||||||||
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Annotation | Half map 1 | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Half map 2
File | emd_4626_half_map_2.map | ||||||||||||
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Annotation | Half map 2 | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : ClpB-DWB-K476C bound to casein in presence of ATPgammaS
Entire | Name: ClpB-DWB-K476C bound to casein in presence of ATPgammaS |
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Components |
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-Supramolecule #1: ClpB-DWB-K476C bound to casein in presence of ATPgammaS
Supramolecule | Name: ClpB-DWB-K476C bound to casein in presence of ATPgammaS type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2 |
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Molecular weight | Theoretical: 570 KDa |
-Supramolecule #2: ClpB-DWB-K476C
Supramolecule | Name: ClpB-DWB-K476C / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1 |
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Source (natural) | Organism: Escherichia coli (E. coli) |
-Supramolecule #3: casein
Supramolecule | Name: casein / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2 |
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Source (natural) | Organism: Bos taurus (cattle) |
-Macromolecule #1: Chaperone protein ClpB
Macromolecule | Name: Chaperone protein ClpB / type: protein_or_peptide / ID: 1 / Number of copies: 6 / Enantiomer: LEVO |
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Source (natural) | Organism: Escherichia coli (E. coli) |
Molecular weight | Theoretical: 95.708133 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: MRLDRLTNKF QLALADAQSL ALGHDNQFIE PLHLMSALLN QEGGSVSPLL TSAGINAGQL RTDINQALNR LPQVEGTGGD VQPSQDLVR VLNLCDKLAQ KRGDNFISSE LFVLAALESR GTLADILKAA GATTANITQA IEQMRGGESV NDQGAEDQRQ A LKKYTIDL ...String: MRLDRLTNKF QLALADAQSL ALGHDNQFIE PLHLMSALLN QEGGSVSPLL TSAGINAGQL RTDINQALNR LPQVEGTGGD VQPSQDLVR VLNLCDKLAQ KRGDNFISSE LFVLAALESR GTLADILKAA GATTANITQA IEQMRGGESV NDQGAEDQRQ A LKKYTIDL TERAEQGKLD PVIGRDEEIR RTIQVLQRRT KNNPVLIGEP GVGKTAIVEG LAQRIINGEV PEGLKGRRVL AL DMGALVA GAKYRGEFEE RLKGVLNDLA KQEGNVILFI DALHTMVGAG KADGAMDAGN MLKPALARGE LHCVGATTLD EYR QYIEKD AALERRFQKV FVAEPSVEDT IAILRGLKER YELHHHVQIT DPAIVAAATL SHRYIADRQL PDKAIDLIDE AASS IRMQI DSKPEELDRL DRRIIQLKLE QQALMKESDE ASKKRLDMLN EELSDKERQY SELEEEWKAE KASLSGTQTI CAELE QAKI AIEQARRVGD LARMSELQYG KIPELEKQLE AATQLEGKTM RLLRNKVTDA EIAEVLARWT GIPVSRMMES EREKLL RME QELHHRVIGQ NEAVDAVSNA IRRSRAGLAD PNRPIGSFLF LGPTGVGKTE LCKALANFMF DSDEAMVRID MSEFMEK HS VSRLVGAPPG YVGYEEGGYL TEAVRRRPYS VILLDAVEKA HPDVFNILLQ VLDDGRLTDG QGRTVDFRNT VVIMTSNL G SDLIQERFGE LDYAHMKELV LGVVSHNFRP EFINRIDEVV VFHPLGEQHI ASIAQIQLKR LYKRLEERGY EIHISDEAL KLLSENGYDP VYGARPLKRA IQQQIENPLA QQILSGELVP GKVIRLEVNE DRIVAVQH UniProtKB: Chaperone protein ClpB |
-Macromolecule #2: casein
Macromolecule | Name: casein / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Bos taurus (cattle) |
Molecular weight | Theoretical: 2.060531 KDa |
Sequence | String: (UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK) |
-Macromolecule #3: PHOSPHOTHIOPHOSPHORIC ACID-ADENYLATE ESTER
Macromolecule | Name: PHOSPHOTHIOPHOSPHORIC ACID-ADENYLATE ESTER / type: ligand / ID: 3 / Number of copies: 10 / Formula: AGS |
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Molecular weight | Theoretical: 523.247 Da |
Chemical component information | ChemComp-AGS: |
-Macromolecule #4: MAGNESIUM ION
Macromolecule | Name: MAGNESIUM ION / type: ligand / ID: 4 / Number of copies: 8 / Formula: MG |
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Molecular weight | Theoretical: 24.305 Da |
-Macromolecule #5: ADENOSINE-5'-DIPHOSPHATE
Macromolecule | Name: ADENOSINE-5'-DIPHOSPHATE / type: ligand / ID: 5 / Number of copies: 1 / Formula: ADP |
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Molecular weight | Theoretical: 427.201 Da |
Chemical component information | ChemComp-ADP: |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 1.6 mg/mL | ||||||||||||||||||
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Buffer | pH: 7.5 Component:
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Grid | Model: Quantifoil, UltrAuFoil / Material: GOLD / Mesh: 300 / Support film - Material: GRAPHENE OXIDE / Support film - topology: CONTINUOUS / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. / Pretreatment - Atmosphere: AIR | ||||||||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 295 K / Instrument: FEI VITROBOT MARK IV |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Specialist optics | Energy filter - Name: GIF Bioquantum |
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Number real images: 6060 / Average electron dose: 1.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 70.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 3.5 µm / Nominal defocus min: 1.5 µm |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |