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- PDB-9skq: Cryo-EM structure of CAK-CDK1-cyclin B1 -

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Entry
Database: PDB / ID: 9skq
TitleCryo-EM structure of CAK-CDK1-cyclin B1
Components
  • (Cyclin-dependent kinase ...) x 2
  • CDK-activating kinase assembly factor MAT1
  • Cyclin-H
  • G2/mitotic-specific cyclin-B1
KeywordsTRANSFERASE / Complex / cell cycle / kinase
Function / homology
Function and homology information


regulation of Schwann cell differentiation / cyclin A1-CDK1 complex / regulation of attachment of mitotic spindle microtubules to kinetochore / pronuclear fusion / response to DDT / cyclin B1-CDK1 complex / positive regulation of mitochondrial ATP synthesis coupled electron transport / Mitotic Prophase / regulation of chromosome condensation / positive regulation of mitotic sister chromatid segregation ...regulation of Schwann cell differentiation / cyclin A1-CDK1 complex / regulation of attachment of mitotic spindle microtubules to kinetochore / pronuclear fusion / response to DDT / cyclin B1-CDK1 complex / positive regulation of mitochondrial ATP synthesis coupled electron transport / Mitotic Prophase / regulation of chromosome condensation / positive regulation of mitotic sister chromatid segregation / histone kinase activity / Golgi disassembly / microtubule cytoskeleton organization involved in mitosis / G2/M DNA replication checkpoint / E2F-enabled inhibition of pre-replication complex formation / ventricular cardiac muscle cell development / Depolymerization of the Nuclear Lamina / positive regulation of attachment of spindle microtubules to kinetochore / MASTL Facilitates Mitotic Progression / positive regulation of mRNA 3'-end processing / regulation of mitotic cell cycle spindle assembly checkpoint / Activation of NIMA Kinases NEK9, NEK6, NEK7 / mitotic nuclear membrane disassembly / : / Phosphorylation of Emi1 / patched binding / RNA polymerase II CTD heptapeptide repeat S5 kinase activity / cyclin A2-CDK1 complex / Transcriptional regulation by RUNX2 / ventricular system development / tissue regeneration / Phosphorylation of the APC/C / Nuclear Pore Complex (NPC) Disassembly / snRNA transcription by RNA polymerase II / CAK-ERCC2 complex / transcription factor TFIIK complex / outer kinetochore / protein localization to kinetochore / Transcription of E2F targets under negative control by p107 (RBL1) and p130 (RBL2) in complex with HDAC1 / adult heart development / Initiation of Nuclear Envelope (NE) Reformation / Polo-like kinase mediated events / Golgi Cisternae Pericentriolar Stack Reorganization / transcription factor TFIIH core complex / transcription factor TFIIH holo complex / cellular response to fatty acid / cyclin-dependent protein serine/threonine kinase activator activity / Condensation of Prometaphase Chromosomes / chromosome condensation / centrosome cycle / digestive tract development / oocyte maturation / [RNA-polymerase]-subunit kinase / RNA Polymerase I Transcription Termination / cyclin-dependent protein serine/threonine kinase regulator activity / response to copper ion / mitotic metaphase chromosome alignment / response to amine / MAPK3 (ERK1) activation / cyclin-dependent protein kinase activity / RNA Pol II CTD phosphorylation and interaction with CE during HIV infection / RNA Pol II CTD phosphorylation and interaction with CE / HIV Transcription Initiation / RNA Polymerase II HIV Promoter Escape / Transcription of the HIV genome / RNA Polymerase II Promoter Escape / RNA Polymerase II Transcription Pre-Initiation And Promoter Opening / RNA Polymerase II Transcription Initiation / RNA Polymerase II Transcription Initiation And Promoter Clearance / Formation of the Early Elongation Complex / Formation of the HIV-1 Early Elongation Complex / G1/S-Specific Transcription / mRNA Capping / mitotic G2 DNA damage checkpoint signaling / microtubule organizing center / Regulation of APC/C activators between G1/S and early anaphase / ubiquitin-like protein ligase binding / RNA Polymerase I Transcription Initiation / regulation of G1/S transition of mitotic cell cycle / regulation of embryonic development / peptidyl-threonine phosphorylation / RNA polymerase II transcribes snRNA genes / protein deubiquitination / positive regulation of G2/M transition of mitotic cell cycle / ATP-dependent activity, acting on DNA / response to mechanical stimulus / response to cadmium ion / cyclin-dependent kinase / cyclin-dependent protein serine/threonine kinase activity / Tat-mediated elongation of the HIV-1 transcript / Formation of HIV-1 elongation complex containing HIV-1 Tat / response to axon injury / Chk1/Chk2(Cds1) mediated inactivation of Cyclin B:Cdk1 complex / Regulation of MITF-M-dependent genes involved in cell cycle and proliferation / Formation of HIV elongation complex in the absence of HIV Tat / Cyclin E associated events during G1/S transition / Cyclin A/B1/B2 associated events during G2/M transition / Nuclear events stimulated by ALK signaling in cancer / Cyclin A:Cdk2-associated events at S phase entry / positive regulation of cardiac muscle cell proliferation
Similarity search - Function
: / CyclinH/Ccl1 / Cyclin-dependent kinase 7 / Cdk-activating kinase assembly factor MAT1/Tfb3 / Cdk-activating kinase assembly factor MAT1, centre / CDK-activating kinase assembly factor MAT1 / Zinc finger, C3HC4 type (RING finger) / Cyclin, C-terminal domain 2 / Cyclin C-terminal domain / : ...: / CyclinH/Ccl1 / Cyclin-dependent kinase 7 / Cdk-activating kinase assembly factor MAT1/Tfb3 / Cdk-activating kinase assembly factor MAT1, centre / CDK-activating kinase assembly factor MAT1 / Zinc finger, C3HC4 type (RING finger) / Cyclin, C-terminal domain 2 / Cyclin C-terminal domain / : / Cyclin, C-terminal domain / : / Cyclins signature. / Cyclin / Cyclin/Cyclin-like subunit Ssn8 / Cyclin, C-terminal domain / Cyclin_C / Ubiquitin interacting motif / Ubiquitin-interacting motif (UIM) domain profile. / Cyclin, N-terminal / Cyclin, N-terminal domain / Cyclin-like / domain present in cyclins, TFIIB and Retinoblastoma / Cyclin-like superfamily / : / Zinc finger, RING-type, conserved site / Zinc finger RING-type signature. / Ring finger / Zinc finger RING-type profile. / Zinc finger, RING-type / Zinc finger, RING/FYVE/PHD-type / Serine/threonine-protein kinase, active site / Serine/Threonine protein kinases active-site signature. / Protein kinase domain / Serine/Threonine protein kinases, catalytic domain / Protein kinase, ATP binding site / Protein kinases ATP-binding region signature. / Protein kinase domain profile. / Protein kinase domain / Protein kinase-like domain superfamily
Similarity search - Domain/homology
PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER / Cyclin-dependent kinase 1 / G2/mitotic-specific cyclin-B1 / Cyclin-dependent kinase 7 / Cyclin-H / CDK-activating kinase assembly factor MAT1
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.4 Å
AuthorsCushing, V.I. / Greber, B.J. / McGeoch, A.J.S. / Feng, J. / Davey, N.E. / Williams, S.L.
Funding support United Kingdom, 3items
OrganizationGrant numberCountry
Medical Research Council (MRC, United Kingdom)MR/V009354/1 United Kingdom
The Institute of Cancer Research (ICR) United Kingdom
Cancer Research UKC68484/A28159 United Kingdom
CitationJournal: To Be Published
Title: Structural basis of T-loop-independent recognition and activation of CDKs by the CDK-activating kinase
Authors: Cushing, V.I. / Greber, B.J. / McGeoch, A.J.S. / Feng, J. / Williams, S.L. / Roumeliotis, T.I. / Choudhary, J.S. / Dan, L.M. / Davey, N.E.
History
DepositionSep 2, 2025Deposition site: PDBE / Processing site: PDBE
Revision 1.0Oct 15, 2025Provider: repository / Type: Initial release
Revision 1.0Oct 15, 2025Data content type: EM metadata / Data content type: EM metadata / Provider: repository / Type: Initial release
Revision 1.0Oct 15, 2025Data content type: Half map / Part number: 1 / Data content type: Half map / Provider: repository / Type: Initial release
Revision 1.0Oct 15, 2025Data content type: Half map / Part number: 2 / Data content type: Half map / Provider: repository / Type: Initial release
Revision 1.0Oct 15, 2025Data content type: Image / Data content type: Image / Provider: repository / Type: Initial release
Revision 1.0Oct 15, 2025Data content type: Mask / Part number: 1 / Data content type: Mask / Provider: repository / Type: Initial release
Revision 1.0Oct 15, 2025Data content type: Primary map / Data content type: Primary map / Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
H: CDK-activating kinase assembly factor MAT1
I: Cyclin-H
J: Cyclin-dependent kinase 7
A: Cyclin-dependent kinase 1
B: G2/mitotic-specific cyclin-B1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)203,9948
Polymers202,9585
Non-polymers1,0373
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

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Protein , 3 types, 3 molecules HIB

#1: Protein CDK-activating kinase assembly factor MAT1 / CDK7/cyclin-H assembly factor / Cyclin-G1-interacting protein / Menage a trois / RING finger ...CDK7/cyclin-H assembly factor / Cyclin-G1-interacting protein / Menage a trois / RING finger protein 66 / RING finger protein MAT1 / p35 / p36


Mass: 38132.340 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: MNAT1, CAP35, MAT1, RNF66 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P51948
#2: Protein Cyclin-H / MO15-associated protein / p34 / p37


Mass: 37721.508 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CCNH / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P51946
#5: Protein G2/mitotic-specific cyclin-B1


Mass: 48800.766 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CCNB1, CCNB / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P14635

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Cyclin-dependent kinase ... , 2 types, 2 molecules JA

#3: Protein Cyclin-dependent kinase 7 / 39 kDa protein kinase / p39 Mo15 / CDK-activating kinase 1 / Cell division protein kinase 7 / ...39 kDa protein kinase / p39 Mo15 / CDK-activating kinase 1 / Cell division protein kinase 7 / Serine/threonine-protein kinase 1 / TFIIH basal transcription factor complex kinase subunit


Mass: 43651.070 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CDK7, CAK, CAK1, CDKN7, MO15, STK1 / Production host: Trichoplusia ni (cabbage looper)
References: UniProt: P50613, cyclin-dependent kinase, [RNA-polymerase]-subunit kinase
#4: Protein Cyclin-dependent kinase 1 / CDK1 / Cell division control protein 2 homolog / Cell division protein kinase 1 / p34 protein kinase


Mass: 34651.902 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CDK1, CDC2, CDC28A, CDKN1, P34CDC2 / Production host: Trichoplusia ni (cabbage looper)
References: UniProt: P06493, cyclin-dependent kinase, [RNA-polymerase]-subunit kinase

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Non-polymers , 2 types, 3 molecules

#6: Chemical ChemComp-ANP / PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER


Mass: 506.196 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C10H17N6O12P3 / Comment: AMP-PNP, energy-carrying molecule analogue*YM
#7: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Mg

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Details

Has ligand of interestN
Has protein modificationY

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Experimental details

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Experiment

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

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Sample preparation

Component
IDNameTypeDetailsEntity IDParent-IDSource
1CAK-CDK1-cyclin B1 complexCOMPLEXCDK-activating kinase (CAK) in complex with CDK1-cyclin B1 in the presence of nucleotide analogue AMP-PNP#1-#50MULTIPLE SOURCES
2CDK-activating kinase (CAK)COMPLEXCAK complex bound to nucleotide analogue AMP-PNP#1-#31RECOMBINANT
3CDK1-cyclin B1COMPLEXCDK1-cyclin B1 complex bound to nucleotide analogue AMP-PNP#4-#51RECOMBINANT
Molecular weight
IDEntity assembly-IDValue (°)Experimental value
110.20 MDaNO
210.12 MDaNO
310.082 MDaNO
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
21Homo sapiens (human)9606
32Homo sapiens (human)9606
43Homo sapiens (human)9606
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-ID
21Trichoplusia ni (cabbage looper)7111
32Trichoplusia ni (cabbage looper)7111
43Trichoplusia ni (cabbage looper)7111
Buffer solutionpH: 7.5
Buffer component
IDConc.NameFormulaBuffer-ID
125 mMHEPES1
2150 mMSodium chlorideNaCl1
310 mMMagnesium chlorideMgCl21
40.5 mMAMP-PNP1
SpecimenConc.: 0.4 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: UltrAuFoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 278 K

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Electron microscopy imaging

MicroscopyModel: TFS GLACIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal magnification: 165000 X / Nominal defocus max: 1900 nm / Nominal defocus min: 500 nm / Cs: 2.7 mm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 60 e/Å2 / Film or detector model: TFS FALCON 4i (4k x 4k) / Num. of grids imaged: 2 / Num. of real images: 14030
EM imaging opticsEnergyfilter name: TFS Selectris

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Processing

EM software
IDNameVersionCategory
1cryoSPARCparticle selection
2EPUimage acquisition
4cryoSPARCCTF correction
5RELION5CTF correction
8UCSF ChimeraXmodel fitting
10RELION5initial Euler assignment
11RELION5final Euler assignment
12RELION4classification
13RELION53D reconstruction
14PHENIX1.21-5207model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 68765 / Num. of class averages: 1 / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT / Space: REAL
Atomic model building
IDPDB-ID 3D fitting-IDAccession codeInitial refinement model-IDSource nameType
19QCV19QCV1PDBexperimental model
25HQ015HQ02PDBexperimental model
RefinementHighest resolution: 3.4 Å
Stereochemistry target values: REAL-SPACE (WEIGHTED MAP SUM AT ATOM CENTERS)
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0029976
ELECTRON MICROSCOPYf_angle_d0.57913514
ELECTRON MICROSCOPYf_dihedral_angle_d4.8021321
ELECTRON MICROSCOPYf_chiral_restr0.0411500
ELECTRON MICROSCOPYf_plane_restr0.0041706

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