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- EMDB-12048: Focused map- CyclinA-CDK2-class. Ubiquitin ligation to F-box prot... -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-12048 | |||||||||
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Title | Focused map- CyclinA-CDK2-class. Ubiquitin ligation to F-box protein substrates by SCF-RBR E3-E3 super-assembly: CUL1-RBX1-SKP1-SKP2-CKSHS1-Cyclin A-CDK2-p27. Transition State 1 | |||||||||
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Function / homology | ![]() cyclin-dependent protein kinase activating kinase regulator activity / regulation of lens fiber cell differentiation / negative regulation of cardiac muscle tissue regeneration / negative regulation of cyclin-dependent protein kinase activity / positive regulation of protein polyubiquitination / autophagic cell death / Parkin-FBXW7-Cul1 ubiquitin ligase complex / FOXO-mediated transcription of cell cycle genes / F-box domain binding / negative regulation of epithelial cell proliferation involved in prostate gland development ...cyclin-dependent protein kinase activating kinase regulator activity / regulation of lens fiber cell differentiation / negative regulation of cardiac muscle tissue regeneration / negative regulation of cyclin-dependent protein kinase activity / positive regulation of protein polyubiquitination / autophagic cell death / Parkin-FBXW7-Cul1 ubiquitin ligase complex / FOXO-mediated transcription of cell cycle genes / F-box domain binding / negative regulation of epithelial cell proliferation involved in prostate gland development / Phosphorylation of proteins involved in the G2/M transition by Cyclin A:Cdc2 complexes / cyclin A2-CDK1 complex / PcG protein complex / cell cycle G1/S phase transition / Aberrant regulation of mitotic exit in cancer due to RB1 defects / cellular response to luteinizing hormone stimulus / regulation of cell cycle G1/S phase transition / regulation of exit from mitosis / mitotic cell cycle phase transition / cullin-RING ubiquitin ligase complex / epithelial cell proliferation involved in prostate gland development / negative regulation of epithelial cell apoptotic process / negative regulation of cyclin-dependent protein serine/threonine kinase activity / negative regulation of phosphorylation / Cul7-RING ubiquitin ligase complex / Transcription of E2F targets under negative control by p107 (RBL1) and p130 (RBL2) in complex with HDAC1 / positive regulation of ubiquitin protein ligase activity / ubiquitin ligase activator activity / Loss of Function of FBXW7 in Cancer and NOTCH1 Signaling / maintenance of protein location in nucleus / cyclin-dependent protein serine/threonine kinase inhibitor activity / cellular response to leptin stimulus / RHO GTPases activate CIT / male pronucleus / female pronucleus / cyclin-dependent protein serine/threonine kinase activator activity / nuclear export / response to glucagon / cellular response to cocaine / AKT phosphorylates targets in the cytosol / positive regulation of intracellular estrogen receptor signaling pathway / cyclin-dependent protein serine/threonine kinase regulator activity / Cul4A-RING E3 ubiquitin ligase complex / epithelial cell apoptotic process / SCF ubiquitin ligase complex / cellular response to insulin-like growth factor stimulus / cellular response to antibiotic / ubiquitin ligase complex scaffold activity / SCF-dependent proteasomal ubiquitin-dependent protein catabolic process / negative regulation of kinase activity / positive regulation of DNA biosynthetic process / regulation of cyclin-dependent protein serine/threonine kinase activity / molecular function inhibitor activity / cochlea development / cellular response to lithium ion / 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 / protein kinase inhibitor activity / positive regulation of heterochromatin formation / p53-Dependent G1 DNA Damage Response / Prolactin receptor signaling / X chromosome / PTK6 Regulates Cell Cycle / protein monoubiquitination / regulation of DNA replication / regulation of anaphase-promoting complex-dependent catabolic process / Constitutive Signaling by AKT1 E17K in Cancer / regulation of G1/S transition of mitotic cell cycle / Defective binding of RB1 mutants to E2F1,(E2F2, E2F3) / cullin family protein binding / negative regulation of vascular associated smooth muscle cell proliferation / centriole replication / Regulation of APC/C activators between G1/S and early anaphase / telomere maintenance in response to DNA damage / centrosome duplication / inner ear development / protein K63-linked ubiquitination / G0 and Early G1 / cellular response to organic cyclic compound / Telomere Extension By Telomerase / negative regulation of mitotic cell cycle / positive regulation of double-strand break repair via homologous recombination / ubiquitin-like ligase-substrate adaptor activity / Activation of the pre-replicative complex / Estrogen-dependent nuclear events downstream of ESR-membrane signaling / cellular response to nitric oxide / response to amino acid / Regulation of MITF-M-dependent genes involved in cell cycle and proliferation / cyclin-dependent kinase / animal organ regeneration / protein K48-linked ubiquitination / cyclin-dependent protein serine/threonine kinase activity Similarity search - Function | |||||||||
Biological species | ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.8 Å | |||||||||
![]() | Horn-Ghetko D / Prabu JR / Schulman BA | |||||||||
Funding support | ![]()
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![]() | ![]() Title: Ubiquitin ligation to F-box protein targets by SCF-RBR E3-E3 super-assembly. Authors: Daniel Horn-Ghetko / David T Krist / J Rajan Prabu / Kheewoong Baek / Monique P C Mulder / Maren Klügel / Daniel C Scott / Huib Ovaa / Gary Kleiger / Brenda A Schulman / ![]() ![]() ![]() Abstract: E3 ligases are typically classified by hallmark domains such as RING and RBR, which are thought to specify unique catalytic mechanisms of ubiquitin transfer to recruited substrates. However, rather ...E3 ligases are typically classified by hallmark domains such as RING and RBR, which are thought to specify unique catalytic mechanisms of ubiquitin transfer to recruited substrates. However, rather than functioning individually, many neddylated cullin-RING E3 ligases (CRLs) and RBR-type E3 ligases in the ARIH family-which together account for nearly half of all ubiquitin ligases in humans-form E3-E3 super-assemblies. Here, by studying CRLs in the SKP1-CUL1-F-box (SCF) family, we show how neddylated SCF ligases and ARIH1 (an RBR-type E3 ligase) co-evolved to ubiquitylate diverse substrates presented on various F-box proteins. We developed activity-based chemical probes that enabled cryo-electron microscopy visualization of steps in E3-E3 ubiquitylation, initiating with ubiquitin linked to the E2 enzyme UBE2L3, then transferred to the catalytic cysteine of ARIH1, and culminating in ubiquitin linkage to a substrate bound to the SCF E3 ligase. The E3-E3 mechanism places the ubiquitin-linked active site of ARIH1 adjacent to substrates bound to F-box proteins (for example, substrates with folded structures or limited length) that are incompatible with previously described conventional RING E3-only mechanisms. The versatile E3-E3 super-assembly may therefore underlie widespread ubiquitylation. | |||||||||
History |
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Structure visualization
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 8.2 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 23.5 KB 23.5 KB | Display Display | ![]() |
FSC (resolution estimation) | ![]() | 11.4 KB | Display | ![]() |
Images | ![]() | 54.3 KB | ||
Masks | ![]() | 125 MB | ![]() | |
Others | ![]() ![]() | 98.4 MB 98.3 MB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 7b5rMC ![]() 7b5lC ![]() 7b5mC ![]() 7b5nC ![]() 7b5sC C: citing same article ( M: atomic model generated by this map |
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Similar structure data |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Voxel size | X=Y=Z: 1.09 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Mask #1
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Projections & Slices |
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Density Histograms |
-Half map: #1
File | emd_12048_half_map_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #2
File | emd_12048_half_map_2.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
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Sample components
-Entire : NEDD8-CUL1-RBX1-SKP1-SKP2-CKSHS1-Cyclin A-CDK2-p27-UBE2L3~Ub~ARIH...
Entire | Name: NEDD8-CUL1-RBX1-SKP1-SKP2-CKSHS1-Cyclin A-CDK2-p27-UBE2L3~Ub~ARIH1. Transition State 1 composite map. |
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Components |
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-Supramolecule #1: NEDD8-CUL1-RBX1-SKP1-SKP2-CKSHS1-Cyclin A-CDK2-p27-UBE2L3~Ub~ARIH...
Supramolecule | Name: NEDD8-CUL1-RBX1-SKP1-SKP2-CKSHS1-Cyclin A-CDK2-p27-UBE2L3~Ub~ARIH1. Transition State 1 composite map. type: complex / ID: 1 / Parent: 0 / Macromolecule list: all |
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Source (natural) | Organism: ![]() |
Recombinant expression | Organism: ![]() ![]() |
Molecular weight | Theoretical: 300 KDa |
-Macromolecule #1: Cullin-1
Macromolecule | Name: Cullin-1 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 89.800367 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MSSTRSQNPH GLKQIGLDQI WDDLRAGIQQ VYTRQSMAKS RYMELYTHVY NYCTSVHQSN QARGAGVPPS KSKKGQTPGG AQFVGLELY KRLKEFLKNY LTNLLKDGED LMDESVLKFY TQQWEDYRFS SKVLNGICAY LNRHWVRREC DEGRKGIYEI Y SLALVTWR ...String: MSSTRSQNPH GLKQIGLDQI WDDLRAGIQQ VYTRQSMAKS RYMELYTHVY NYCTSVHQSN QARGAGVPPS KSKKGQTPGG AQFVGLELY KRLKEFLKNY LTNLLKDGED LMDESVLKFY TQQWEDYRFS SKVLNGICAY LNRHWVRREC DEGRKGIYEI Y SLALVTWR DCLFRPLNKQ VTNAVLKLIE KERNGETINT RLISGVVQSY VELGLNEDDA FAKGPTLTVY KESFESQFLA DT ERFYTRE STEFLQQNPV TEYMKKAEAR LLEEQRRVQV YLHESTQDEL ARKCEQVLIE KHLEIFHTEF QNLLDADKNE DLG RMYNLV SRIQDGLGEL KKLLETHIHN QGLAAIEKCG EAALNDPKMY VQTVLDVHKK YNALVMSAFN NDAGFVAALD KACG RFINN NAVTKMAQSS SKSPELLARY CDSLLKKSSK NPEEAELEDT LNQVMVVFKY IEDKDVFQKF YAKMLAKRLV HQNSA SDDA EASMISKLKQ ACGFEYTSKL QRMFQDIGVS KDLNEQFKKH LTNSEPLDLD FSIQVLSSGS WPFQQSCTFA LPSELE RSY QRFTAFYASR HSGRKLTWLY QLSKGELVTN CFKNRYTLQA STFQMAILLQ YNTEDAYTVQ QLTDSTQIKM DILAQVL QI LLKSKLLVLE DENANVDEVE LKPDTLIKLY LGYKNKKLRV NINVPMKTEQ KQEQETTHKN IEEDRKLLIQ AAIVRIMK M RKVLKHQQLL GEVLTQLSSR FKPRVPVIKK CIDILIEKEY LERVDGEKDT YSYLA |
-Macromolecule #2: S-phase kinase-associated protein 2
Macromolecule | Name: S-phase kinase-associated protein 2 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 47.817785 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MHRKHLQEIP DLSSNVATSF TWGWDSSKTS ELLSGMGVSA LEKEEPDSEN IPQELLSNLG HPESPPRKRL KSKGSDKDFV IVRRPKLNR ENFPGVSWDS LPDELLLGIF SCLCLPELLK VSGVCKRWYR LASDESLWQT LDLTGKNLHP DVTGRLLSQG V IAFRCPRS ...String: MHRKHLQEIP DLSSNVATSF TWGWDSSKTS ELLSGMGVSA LEKEEPDSEN IPQELLSNLG HPESPPRKRL KSKGSDKDFV IVRRPKLNR ENFPGVSWDS LPDELLLGIF SCLCLPELLK VSGVCKRWYR LASDESLWQT LDLTGKNLHP DVTGRLLSQG V IAFRCPRS FMDQPLAEHF SPFRVQHMDL SNSVIEVSTL HGILSQCSKL QNLSLEGLRL SDPIVNTLAK NSNLVRLNLS GC SGFSEFA LQTLLSSCSR LDELNLSWCF DFTEKHVQVA VAHVSETITQ LNLSGYRKNL QKSDLSTLVR RCPNLVHLDL SDS VMLKND CFQEFFQLNY LQHLSLSRCY DIIPETLLEL GEIPTLKTLQ VFGIVPDGTL QLLKEALPHL QINCSHFTTI ARPT IGNKK NQEIWGIKCR LTLQKPSCL |
-Macromolecule #3: Cyclin-dependent kinases regulatory subunit 1
Macromolecule | Name: Cyclin-dependent kinases regulatory subunit 1 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 9.679211 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MSHKQIYYSD KYDDEEFEYR HVMLPKDIAK LVPKTHLMSE SEWRNLGVQQ SQGWVHYMIH EPEPHILLFR RPLPKKPKK |
-Macromolecule #4: S-phase kinase-associated protein 1
Macromolecule | Name: S-phase kinase-associated protein 1 / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 18.679965 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MPSIKLQSSD GEIFEVDVEI AKQSVTIKTM LEDLGMDDEG DDDPVPLPNV NAAILKKVIQ WCTHHKDDPP PPEDDENKEK RTDDIPVWD QEFLKVDQGT LFELILAANY LDIKGLLDVT CKTVANMIKG KTPEEIRKTF NIKNDFTEEE EAQVRKENQW C EEK |
-Macromolecule #5: Cyclin-dependent kinase 2
Macromolecule | Name: Cyclin-dependent kinase 2 / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO / EC number: cyclin-dependent kinase |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 34.056469 KDa |
Recombinant expression | Organism: ![]() ![]() |
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 L |
-Macromolecule #6: Cyclin-A2
Macromolecule | Name: Cyclin-A2 / type: protein_or_peptide / ID: 6 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 48.609574 KDa |
Recombinant expression | Organism: ![]() ![]() |
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 |
-Macromolecule #7: Cyclin-dependent kinase inhibitor 1B
Macromolecule | Name: Cyclin-dependent kinase inhibitor 1B / type: protein_or_peptide / ID: 7 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 22.188303 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MSNVRVSNGS PSLERMDARQ AEHPKPSACR NLFGPVDHEE LTRDLEKHCR DMEEASQRKW NFDFQNHKPL EGKYEWQEVE KGSLPEFYY RPPRPPKGAC KVPAQESQDV SGSRPAAPLI GAPANSEDTH LVDPKTDPSD SQTGLAEQCA GIRKRPATDD S STQNKRAN ...String: MSNVRVSNGS PSLERMDARQ AEHPKPSACR NLFGPVDHEE LTRDLEKHCR DMEEASQRKW NFDFQNHKPL EGKYEWQEVE KGSLPEFYY RPPRPPKGAC KVPAQESQDV SGSRPAAPLI GAPANSEDTH LVDPKTDPSD SQTGLAEQCA GIRKRPATDD S STQNKRAN RTEENVSDGS PNAGSVEQ(TPO)P KKPGLRRRQT |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Buffer | pH: 7.8 |
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Vitrification | Cryogen name: ETHANE |
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Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 70.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |