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- EMDB-19408: Cryo-EM structure of CDK2-cyclin A in complex with CDC25A -

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Basic information

Entry
Database: EMDB / ID: EMD-19408
TitleCryo-EM structure of CDK2-cyclin A in complex with CDC25A
Map data
Sample
  • Complex: Trimeric complex of CDK2-cyclin A-CDC25A
    • Protein or peptide: Cyclin-dependent kinase 2
    • Protein or peptide: Cyclin-A2
    • Protein or peptide: M-phase inducer phosphatase 1
Keywordscell-cycle / CDK / phosphatase / cancer / CELL CYCLE
Function / homology
Function and homology information


positive regulation of G2/MI transition of meiotic cell cycle / cell cycle G1/S phase transition / Polo-like kinase mediated events / Transcription of E2F targets under negative control by DREAM complex / regulation of cyclin-dependent protein serine/threonine kinase activity / cyclin-dependent protein serine/threonine kinase regulator activity / Deregulated CDK5 triggers multiple neurodegenerative pathways in Alzheimer's disease models / cyclin A1-CDK2 complex / cyclin E2-CDK2 complex / cyclin E1-CDK2 complex ...positive regulation of G2/MI transition of meiotic cell cycle / cell cycle G1/S phase transition / Polo-like kinase mediated events / Transcription of E2F targets under negative control by DREAM complex / regulation of cyclin-dependent protein serine/threonine kinase activity / cyclin-dependent protein serine/threonine kinase regulator activity / Deregulated CDK5 triggers multiple neurodegenerative pathways in Alzheimer's disease models / cyclin A1-CDK2 complex / cyclin E2-CDK2 complex / cyclin E1-CDK2 complex / cyclin A2-CDK2 complex / positive regulation of DNA-templated DNA replication initiation / cyclin-dependent protein kinase activity / G2 Phase / Y chromosome / Phosphorylation of proteins involved in G1/S transition by active Cyclin E:Cdk2 complexes / positive regulation of heterochromatin formation / p53-Dependent G1 DNA Damage Response / X chromosome / PTK6 Regulates Cell Cycle / regulation of anaphase-promoting complex-dependent catabolic process / Defective binding of RB1 mutants to E2F1,(E2F2, E2F3) / microtubule organizing center / centriole replication / Regulation of APC/C activators between G1/S and early anaphase / telomere maintenance in response to DNA damage / regulation of DNA replication / centrosome duplication / phosphoprotein phosphatase activity / Telomere Extension By Telomerase / G0 and Early G1 / Activation of the pre-replicative complex / cyclin-dependent kinase / cyclin-dependent protein serine/threonine kinase activity / TP53 Regulates Transcription of Genes Involved in G1 Cell Cycle Arrest / Regulation of MITF-M-dependent genes involved in cell cycle and proliferation / Cajal body / Activation of ATR in response to replication stress / Cyclin E associated events during G1/S transition / Cyclin A/B1/B2 associated events during G2/M transition / Cyclin A:Cdk2-associated events at S phase entry / cyclin-dependent protein kinase holoenzyme complex / condensed chromosome / regulation of G2/M transition of mitotic cell cycle / mitotic G1 DNA damage checkpoint signaling / positive regulation of G2/M transition of mitotic cell cycle / protein tyrosine phosphatase activity / protein-tyrosine-phosphatase / protein tyrosine phosphatase activity, metal-dependent / histone H2AXY142 phosphatase activity / non-membrane spanning protein tyrosine phosphatase activity / cellular response to nitric oxide / post-translational protein modification / cyclin binding / regulation of mitotic cell cycle / male germ cell nucleus / positive regulation of DNA replication / meiotic cell cycle / Ubiquitin Mediated Degradation of Phosphorylated Cdc25A / response to radiation / potassium ion transport / DNA Damage/Telomere Stress Induced Senescence / Meiotic recombination / CDK-mediated phosphorylation and removal of Cdc6 / SCF(Skp2)-mediated degradation of p27/p21 / G1/S transition of mitotic cell cycle / Transcriptional regulation of granulopoiesis / Orc1 removal from chromatin / G2/M transition of mitotic cell cycle / Cyclin D associated events in G1 / cellular senescence / Regulation of TP53 Degradation / nuclear envelope / peptidyl-serine phosphorylation / protein-folding chaperone binding / Factors involved in megakaryocyte development and platelet production / Processing of DNA double-strand break ends / regulation of gene expression / Senescence-Associated Secretory Phenotype (SASP) / Regulation of TP53 Activity through Phosphorylation / transcription regulator complex / Ras protein signal transduction / chromosome, telomeric region / cell population proliferation / DNA replication / endosome / Ub-specific processing proteases / protein phosphorylation / chromatin remodeling / protein domain specific binding / cell division / protein serine kinase activity / DNA repair / protein serine/threonine kinase activity / DNA-templated transcription / positive regulation of cell population proliferation / centrosome / protein kinase binding / positive regulation of DNA-templated transcription / negative regulation of transcription by RNA polymerase II
Similarity search - Function
M-phase inducer phosphatase / M-phase inducer phosphatase / Cyclin-A, N-terminal APC/C binding region / Cyclin-A N-terminal APC/C binding region / Rhodanese Homology Domain / : / Rhodanese-like domain / Rhodanese domain profile. / Rhodanese-like domain superfamily / Cyclin, C-terminal domain ...M-phase inducer phosphatase / M-phase inducer phosphatase / Cyclin-A, N-terminal APC/C binding region / Cyclin-A N-terminal APC/C binding region / Rhodanese Homology Domain / : / Rhodanese-like domain / Rhodanese domain profile. / Rhodanese-like domain superfamily / Cyclin, C-terminal domain / : / Cyclins signature. / Cyclin / Rhodanese-like domain / Cyclin, C-terminal domain / Cyclin_C / Cyclin, N-terminal / Cyclin, N-terminal domain / Cyclin-like / domain present in cyclins, TFIIB and Retinoblastoma / Cyclin-like superfamily / : / 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
Cyclin-dependent kinase 2 / Cyclin-A2 / M-phase inducer phosphatase 1
Similarity search - Component
Biological speciesHomo sapiens (human) / Bos taurus (domestic cattle)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.7 Å
AuthorsRowland RJ / Noble MEM / Endicott JA
Funding support United Kingdom, 2 items
OrganizationGrant numberCountry
Medical Research Council (MRC, United Kingdom)MR/V029142/1 United Kingdom
Medical Research Council (MRC, United Kingdom)MR/N009738/1 United Kingdom
CitationJournal: Nat Commun / Year: 2024
Title: Cryo-EM structure of the CDK2-cyclin A-CDC25A complex.
Authors: Rhianna J Rowland / Svitlana Korolchuk / Marco Salamina / Natalie J Tatum / James R Ault / Sam Hart / Johan P Turkenburg / James N Blaza / Martin E M Noble / Jane A Endicott /
Abstract: The cell division cycle 25 phosphatases CDC25A, B and C regulate cell cycle transitions by dephosphorylating residues in the conserved glycine-rich loop of CDKs to activate their activity. Here, we ...The cell division cycle 25 phosphatases CDC25A, B and C regulate cell cycle transitions by dephosphorylating residues in the conserved glycine-rich loop of CDKs to activate their activity. Here, we present the cryo-EM structure of CDK2-cyclin A in complex with CDC25A at 2.7 Å resolution, providing a detailed structural analysis of the overall complex architecture and key protein-protein interactions that underpin this 86 kDa complex. We further identify a CDC25A C-terminal helix that is critical for complex formation. Sequence conservation analysis suggests CDK1/2-cyclin A, CDK1-cyclin B and CDK2/3-cyclin E are suitable binding partners for CDC25A, whilst CDK4/6-cyclin D complexes appear unlikely substrates. A comparative structural analysis of CDK-containing complexes also confirms the functional importance of the conserved CDK1/2 GDSEID motif. This structure improves our understanding of the roles of CDC25 phosphatases in CDK regulation and may inform the development of CDC25-targeting anticancer strategies.
History
DepositionJan 12, 2024-
Header (metadata) releaseMay 29, 2024-
Map releaseMay 29, 2024-
UpdateNov 13, 2024-
Current statusNov 13, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_19408.png

Downloads & links

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Map

FileDownload / File: emd_19408.map.gz / Format: CCP4 / Size: 91.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

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AxesZ (Sec.)Y (Row.)X (Col.)
0.83 Å/pix.
x 288 pix.
= 237.6 Å		0.83 Å/pix.
x 288 pix.
= 237.6 Å		0.83 Å/pix.
x 288 pix.
= 237.6 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.825 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.08
Minimum - Maximum-0.6334735 - 0.9438938
Average (Standard dev.)0.00021430186 (±0.017006245)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions288288288
Spacing288288288
CellA=B=C: 237.59999 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_19408_msk_1.map
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Half map: #2

Fileemd_19408_half_map_1.map
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Half map: #1

Fileemd_19408_half_map_2.map
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Sample components

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Entire : Trimeric complex of CDK2-cyclin A-CDC25A

EntireName: Trimeric complex of CDK2-cyclin A-CDC25A
Components
  • Complex: Trimeric complex of CDK2-cyclin A-CDC25A
    • Protein or peptide: Cyclin-dependent kinase 2
    • Protein or peptide: Cyclin-A2
    • Protein or peptide: M-phase inducer phosphatase 1

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Supramolecule #1: Trimeric complex of CDK2-cyclin A-CDC25A

SupramoleculeName: Trimeric complex of CDK2-cyclin A-CDC25A / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 86 KDa

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Macromolecule #1: Cyclin-dependent kinase 2

MacromoleculeName: Cyclin-dependent kinase 2 / type: protein_or_peptide / ID: 1
Details: Human cyclin dependant protein kinase 2 (CDK2) phosphorylated at Tyr15 and Thr160
Number of copies: 1 / Enantiomer: LEVO / EC number: cyclin-dependent kinase
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 34.136449 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MENFQKVEKI GEGT(PTR)GVVYK ARNKLTGEVV ALKKIRLDTE TEGVPSTAIR EISLLKELNH PNIVKLLDVI HTENKL YLV FEFLHQDLKK FMDASALTGI PLPLIKSYLF QLLQGLAFCH SHRVLHRDLK PQNLLINTEG AIKLADFGLA RAFGVPV RT Y(TPO) ...String:
MENFQKVEKI GEGT(PTR)GVVYK ARNKLTGEVV ALKKIRLDTE TEGVPSTAIR EISLLKELNH PNIVKLLDVI HTENKL YLV FEFLHQDLKK FMDASALTGI PLPLIKSYLF QLLQGLAFCH SHRVLHRDLK PQNLLINTEG AIKLADFGLA RAFGVPV RT Y(TPO)HEVVTLWY RAPEILLGCK YYSTAVDIWS LGCIFAEMVT RRALFPGDSE IDQLFRIFRT LGTPDEVVWP GVTS MPDYK PSFPKWARQD FSKVVPPLDE DGRSLLSQML HYDPNKRISA KAALAHPFFQ DVTKPVPHLR L

UniProtKB: Cyclin-dependent kinase 2

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Macromolecule #2: Cyclin-A2

MacromoleculeName: Cyclin-A2 / type: protein_or_peptide / ID: 2 / Details: Bovine cyclin A2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Bos taurus (domestic cattle)
Molecular weightTheoretical: 30.119799 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MGVNEVPDYH EDIHTYLREM EVKCKPKVGY MKKQPDITNS MRAILVDWLV EVGEEYKLQN ETLHLAVNYI DRFLSSMSVL RGKLQLVGT AAMLLASKFE EIYPPEVAEF VYITDDTYTK KQVLRMEHLV LKVLAFDLAA PTINQFLTQY FLHQQPANCK V ESLAMFLG ...String:
MGVNEVPDYH EDIHTYLREM EVKCKPKVGY MKKQPDITNS MRAILVDWLV EVGEEYKLQN ETLHLAVNYI DRFLSSMSVL RGKLQLVGT AAMLLASKFE EIYPPEVAEF VYITDDTYTK KQVLRMEHLV LKVLAFDLAA PTINQFLTQY FLHQQPANCK V ESLAMFLG ELSLIDADPY LKYLPSVIAA AAFHLALYTV TGQSWPESLV QKTGYTLETL KPCLLDLHQT YLRAPQHAQQ SI REKYKNS KYHGVSLLNP PETLNV

UniProtKB: Cyclin-A2

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Macromolecule #3: M-phase inducer phosphatase 1

MacromoleculeName: M-phase inducer phosphatase 1 / type: protein_or_peptide / ID: 3
Details: M-phase inducer phosphatase 1 (CDC25A) catalytic domain
Number of copies: 1 / Enantiomer: LEVO / EC number: protein-tyrosine-phosphatase
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 22.533123 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString:
SMVIGDFSKG YLFHTVAGKH QDLKYISPEI MASVLNGKFA NLIKEFVIID CRYPYEYEGG HIKGAVNLHM EEEVEDFLLK KPIVPTDGK RVIVVFHSEF SSERGPRMCR YVRERDRLGN EYPKLHYPEL YVLKGGYKEF FMKCQSYCEP PSYRPMHHED F KEDLKKFR TKSRTWAGEK SKREMYSRLK KL

UniProtKB: M-phase inducer phosphatase 1

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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

Concentration1.8 mg/mL
BufferpH: 7.4
Component:
ConcentrationFormulaName
50.0 mMHEPESHEPES
200.0 mMNaClSalt
1.0 mMDTTDTT

Details: 50 mM HEPES, 200 mM NaCl, 1 mM DTT, pH 7.4
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 0.026000000000000002 kPa
Details: Grids were glow discharged for 1min at 20 mAmp/0.26 mBar
VitrificationCryogen name: ETHANE / Chamber humidity: 80 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeTFS KRIOS
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average exposure time: 1.92 sec. / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.6 µm / Nominal magnification: 150000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 4981705
Startup modelType of model: NONE
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 2.7 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 670830
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC / Software - details: Ab initio
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC / Software - details: non-uniform refinement
Final 3D classificationNumber classes: 6 / Software - Name: cryoSPARC
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelChain - Source name: Other / Chain - Initial model type: in silico model / Details: ModelAngelo
DetailsA model was generated using ModelAngelo. Initial local fitting was performed in ChimerX followed by real space refinement in Phenix and manual fitting in coot.
RefinementSpace: REAL / Protocol: AB INITIO MODEL / Overall B value: 144
Output model

PDB-8roz:
Cryo-EM structure of CDK2-cyclin A in complex with CDC25A

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