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Yorodumi- EMDB-16344: Cryo-EM structure of CDK2-CyclinA in complex with p27 from the SC... -
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-Basic information
Entry | Database: EMDB / ID: EMD-16344 | |||||||||
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Title | Cryo-EM structure of CDK2-CyclinA in complex with p27 from the SCFSKP2 E3 ligase Complex | |||||||||
Map data | full map | |||||||||
Sample |
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Keywords | cell cycle / cyclin-dependent kinase / signalling / ubiquitination | |||||||||
Function / homology | Function and homology information Phosphorylation of proteins involved in the G2/M transition by Cyclin A:Cdc2 complexes / cyclin A2-CDK1 complex / cell cycle G1/S phase transition / cellular response to luteinizing hormone stimulus / mitotic cell cycle phase transition / Transcription of E2F targets under negative control by p107 (RBL1) and p130 (RBL2) in complex with HDAC1 / cyclin-dependent protein serine/threonine kinase inhibitor activity / cellular response to leptin stimulus / male pronucleus / female pronucleus ...Phosphorylation of proteins involved in the G2/M transition by Cyclin A:Cdc2 complexes / cyclin A2-CDK1 complex / cell cycle G1/S phase transition / cellular response to luteinizing hormone stimulus / mitotic cell cycle phase transition / Transcription of E2F targets under negative control by p107 (RBL1) and p130 (RBL2) in complex with HDAC1 / cyclin-dependent protein serine/threonine kinase inhibitor activity / cellular response to leptin stimulus / male pronucleus / female pronucleus / cellular response to cocaine / response to glucagon / cyclin-dependent protein serine/threonine kinase regulator activity / cellular response to insulin-like growth factor stimulus / positive regulation of DNA biosynthetic process / cochlea development / cyclin A1-CDK2 complex / cyclin E2-CDK2 complex / cyclin E1-CDK2 complex / cellular response to platelet-derived growth factor stimulus / cyclin A2-CDK2 complex / positive regulation of DNA-templated DNA replication initiation / G2 Phase / cyclin-dependent protein kinase activity / 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) / regulation of DNA replication / centriole replication / Regulation of APC/C activators between G1/S and early anaphase / centrosome duplication / Telomere Extension By Telomerase / G0 and Early G1 / Activation of the pre-replicative complex / cyclin-dependent protein kinase holoenzyme complex / cellular response to nitric oxide / cyclin-dependent kinase / Cajal body / animal organ regeneration / cyclin-dependent protein serine/threonine kinase activity / TP53 Regulates Transcription of Genes Involved in G1 Cell Cycle Arrest / 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 / condensed chromosome / mitotic G1 DNA damage checkpoint signaling / regulation of mitotic cell cycle / regulation of G2/M transition of mitotic cell cycle / cyclin binding / post-translational protein modification / meiotic cell cycle / positive regulation of DNA replication / response to organic substance / male germ cell nucleus / cellular response to estradiol stimulus / Cdc20:Phospho-APC/C mediated degradation of Cyclin A / G1/S transition of mitotic cell cycle / potassium ion transport / DNA Damage/Telomere Stress Induced Senescence / CDK-mediated phosphorylation and removal of Cdc6 / Meiotic recombination / SCF(Skp2)-mediated degradation of p27/p21 / Orc1 removal from chromatin / Transcriptional regulation of granulopoiesis / Cyclin D associated events in G1 / G2/M transition of mitotic cell cycle / positive regulation of fibroblast proliferation / cellular senescence / Regulation of TP53 Degradation / nuclear envelope / Factors involved in megakaryocyte development and platelet production / Processing of DNA double-strand break ends / cellular response to hypoxia / Senescence-Associated Secretory Phenotype (SASP) / regulation of gene expression / peptidyl-serine phosphorylation / Ras protein signal transduction / Regulation of TP53 Activity through Phosphorylation / transcription regulator complex / DNA replication / chromosome, telomeric region / regulation of cell cycle / Ub-specific processing proteases / endosome / chromatin remodeling / cell division / protein domain specific binding / protein phosphorylation / protein serine kinase activity / DNA repair / protein serine/threonine kinase activity / centrosome / DNA-templated transcription / positive regulation of cell population proliferation Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.5 Å | |||||||||
Authors | Rowland RJ / Salamina M / Endicott JA / Noble ME | |||||||||
Funding support | United Kingdom, 1 items
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Citation | Journal: Sci Rep / Year: 2023 Title: Cryo-EM structure of SKP1-SKP2-CKS1 in complex with CDK2-cyclin A-p27KIP1. Authors: Rhianna J Rowland / Richard Heath / Daniel Maskell / Rebecca F Thompson / Neil A Ranson / James N Blaza / Jane A Endicott / Martin E M Noble / Marco Salamina / Abstract: p27KIP1 (cyclin-dependent kinase inhibitor 1B, p27) is a member of the CIP/KIP family of CDK (cyclin dependent kinase) regulators that inhibit cell cycle CDKs. p27 phosphorylation by CDK1/2, signals ...p27KIP1 (cyclin-dependent kinase inhibitor 1B, p27) is a member of the CIP/KIP family of CDK (cyclin dependent kinase) regulators that inhibit cell cycle CDKs. p27 phosphorylation by CDK1/2, signals its recruitment to the SCF (S-phase kinase associated protein 1 (SKP1)-cullin-SKP2) E3 ubiquitin ligase complex for proteasomal degradation. The nature of p27 binding to SKP2 and CKS1 was revealed by the SKP1-SKP2-CKS1-p27 phosphopeptide crystal structure. Subsequently, a model for the hexameric CDK2-cyclin A-CKS1-p27-SKP1-SKP2 complex was proposed by overlaying an independently determined CDK2-cyclin A-p27 structure. Here we describe the experimentally determined structure of the isolated CDK2-cyclin A-CKS1-p27-SKP1-SKP2 complex at 3.4 Å global resolution using cryogenic electron microscopy. This structure supports previous analysis in which p27 was found to be structurally dynamic, transitioning from disordered to nascent secondary structure on target binding. We employed 3D variability analysis to further explore the conformational space of the hexameric complex and uncovered a previously unidentified hinge motion centred on CKS1. This flexibility gives rise to open and closed conformations of the hexameric complex that we propose may contribute to p27 regulation by facilitating recognition with SCF. This 3D variability analysis further informed particle subtraction and local refinement approaches to enhance the local resolution of the complex. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_16344.map.gz | 97.2 MB | EMDB map data format | |
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Header (meta data) | emd-16344-v30.xml emd-16344.xml | 20.3 KB 20.3 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_16344_fsc.xml | 9.9 KB | Display | FSC data file |
Images | emd_16344.png | 124.9 KB | ||
Masks | emd_16344_msk_1.map | 103 MB | Mask map | |
Others | emd_16344_half_map_1.map.gz emd_16344_half_map_2.map.gz | 95.5 MB 95.5 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-16344 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-16344 | HTTPS FTP |
-Related structure data
Related structure data | 8bzoMC 8byaC 8bylC C: citing same article (ref.) M: atomic model generated by this map |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_16344.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||
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Annotation | full map | ||||||||||||||||||||
Voxel size | X=Y=Z: 1.07 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Mask #1
File | emd_16344_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: half map A
File | emd_16344_half_map_1.map | ||||||||||||
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Annotation | half map A | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: half map B
File | emd_16344_half_map_2.map | ||||||||||||
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Annotation | half map B | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : Complex of CDK2-CyclinA-p27(kip1)from the SCFSKP2 E3 ligase complex
Entire | Name: Complex of CDK2-CyclinA-p27(kip1)from the SCFSKP2 E3 ligase complex |
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Components |
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-Supramolecule #1: Complex of CDK2-CyclinA-p27(kip1)from the SCFSKP2 E3 ligase complex
Supramolecule | Name: Complex of CDK2-CyclinA-p27(kip1)from the SCFSKP2 E3 ligase complex type: complex / ID: 1 / Parent: 0 / Macromolecule list: all |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 74 KDa |
-Macromolecule #1: Cyclin-dependent kinase 2
Macromolecule | Name: Cyclin-dependent kinase 2 / type: protein_or_peptide / ID: 1 / Details: Thr160 phosphorylated CDK2 / Number of copies: 1 / Enantiomer: LEVO / EC number: cyclin-dependent kinase |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 33.943312 KDa |
Recombinant expression | Organism: Escherichia coli BL21(DE3) (bacteria) |
Sequence | String: MENFQKVEKI GEGTYGVVYK ARNKLTGEVV ALKKIRLDTE TEGVPSTAIR EISLLKELNH PNIVKLLDVI HTENKLYLVF EFLHQDLKK FMDASALTGI PLPLIKSYLF QLLQGLAFCH SHRVLHRDLK PQNLLINTEG AIKLADFGLA RAFGVPVRTY (TPO)HEVVTLWY ...String: MENFQKVEKI GEGTYGVVYK ARNKLTGEVV ALKKIRLDTE TEGVPSTAIR EISLLKELNH PNIVKLLDVI HTENKLYLVF EFLHQDLKK FMDASALTGI PLPLIKSYLF QLLQGLAFCH SHRVLHRDLK PQNLLINTEG AIKLADFGLA RAFGVPVRTY (TPO)HEVVTLWY RAPEILLGCK YYSTAVDIWS LGCIFAEMVT RRALFPGDSE IDQLFRIFRT LGTPDEVVWP GVTSMPD YK PSFPKWARQD FSKVVPPLDE DGRSLLSQML HYDPNKRISA KAALAHPFFQ DVTKPVPHLR UniProtKB: Cyclin-dependent kinase 2 |
-Macromolecule #2: Cyclin-A2
Macromolecule | Name: Cyclin-A2 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 48.609574 KDa |
Recombinant expression | Organism: Escherichia coli BL21(DE3) (bacteria) |
Sequence | String: MLGNSAPGPA TREAGSALLA LQQTALQEDQ ENINPEKAAP VQQPRTRAAL AVLKSGNPRG LAQQQRPKTR RVAPLKDLPV NDEHVTVPP WKANSKQPAF TIHVDEAEKE AQKKPAESQK IEREDALAFN SAISLPGPRK PLVPLDYPMD GSFESPHTMD M SIILEDEK ...String: MLGNSAPGPA TREAGSALLA LQQTALQEDQ ENINPEKAAP VQQPRTRAAL AVLKSGNPRG LAQQQRPKTR RVAPLKDLPV NDEHVTVPP WKANSKQPAF TIHVDEAEKE AQKKPAESQK IEREDALAFN SAISLPGPRK PLVPLDYPMD GSFESPHTMD M SIILEDEK PVSVNEVPDY HEDIHTYLRE MEVKCKPKVG YMKKQPDITN SMRAILVDWL VEVGEEYKLQ NETLHLAVNY ID RFLSSMS VLRGKLQLVG TAAMLLASKF EEIYPPEVAE FVYITDDTYT KKQVLRMEHL VLKVLTFDLA APTVNQFLTQ YFL HQQPAN CKVESLAMFL GELSLIDADP YLKYLPSVIA GAAFHLALYT VTGQSWPESL IRKTGYTLES LKPCLMDLHQ TYLK APQHA QQSIREKYKN SKYHGVSLLN PPETLNL UniProtKB: Cyclin-A2 |
-Macromolecule #3: Cyclin-dependent kinase inhibitor 1B
Macromolecule | Name: Cyclin-dependent kinase inhibitor 1B / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 17.678531 KDa |
Recombinant expression | Organism: Escherichia coli BL21(DE3) (bacteria) |
Sequence | String: MSNVRVSNGS PSLERMDARQ AEHPKPSACR NLFGPVDHEE LTRDLEKHCR DMEEASQRKW NFDFQNHKPL EGKYEWQEVE KGSLPEFYY RPPRPPKGAC KVPAQESQDG SGSRPAAPLI GAPANSEDTH LVDPKTDPSD SQTGLAEQCA GIRKRPATD UniProtKB: Cyclin-dependent kinase inhibitor 1B |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.2 mg/mL |
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Buffer | pH: 7.8 |
Grid | Model: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. / Pretreatment - Atmosphere: OTHER |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 278.15 K / Instrument: FEI VITROBOT MARK IV |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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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 FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 130000 |
Sample stage | Cooling holder cryogen: NITROGEN |
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Average exposure time: 9.0 sec. / Average electron dose: 65.0 e/Å2 |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
-Atomic model buiding 1
Details | Initial fitting was performed in chimera followed by real space refinement in Phenix |
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Refinement | Space: REAL / Protocol: RIGID BODY FIT / Overall B value: 121 |
Output model | PDB-8bzo: |