EMDB-18221: Structure of the hexameric CUL9-RBX1 complex with deletion of CUL...

Basic information

Entry

Database: EMDB / ID: EMD-18221

• Title: Structure of the hexameric CUL9-RBX1 complex with deletion of CUL9 DOC domain
• Map data: postprocess map

Sample

• Complex: Structure of the hexameric CUL9-RBX1 complex with deletion of CUL9 DOC domain
  • Protein or peptide: Cullin-9
  • Protein or peptide: E3 ubiquitin-protein ligase RBX1

• Keywords: Cullin-RING RBR E3 Ligase / LIGASE

Function / homology

Function:

negative regulation of beige fat cell differentiation / cullin-RING-type E3 NEDD8 transferase / NEDD8 transferase activity / negative regulation of mitophagy / cullin-RING ubiquitin ligase complex / regulation of xenophagy / Loss of Function of FBXW7 in Cancer and NOTCH1 Signaling / cellular response to chemical stress / Cul7-RING ubiquitin ligase complex / regulation of cell cycle process / neural crest cell differentiation / RNA polymerase II transcription initiation surveillance / positive regulation of protein autoubiquitination / protein neddylation / regulation of BMP signaling pathway / NEDD8 ligase activity / regulation of mitophagy / negative regulation of response to oxidative stress / regulation of centrosome duplication / regulation of mitotic nuclear division / protein K27-linked ubiquitination / VCB complex / Cul5-RING ubiquitin ligase complex / regulation of TOR signaling / ubiquitin-ubiquitin ligase activity / ubiquitin-dependent protein catabolic process via the C-end degron rule pathway / Cul2-RING ubiquitin ligase complex / SCF ubiquitin ligase complex / negative regulation of DNA-templated DNA replication / regulation of mitotic cytokinesis / Cul3-RING ubiquitin ligase complex / regulation of DNA damage checkpoint / negative regulation of type I interferon production / regulation of miRNA-mediated gene silencing / regulation of natural killer cell activation / SCF-dependent proteasomal ubiquitin-dependent protein catabolic process / Prolactin receptor signaling / regulation of cell cycle phase transition / Cul4A-RING E3 ubiquitin ligase complex / Cul4-RING E3 ubiquitin ligase complex / regulation of stem cell population maintenance / Cul4B-RING E3 ubiquitin ligase complex / ubiquitin ligase complex scaffold activity / negative regulation of adipose tissue development / regulation of cellular response to stress / limb development / protein monoubiquitination / cullin family protein binding / regulation of DNA-templated DNA replication initiation / centrosome duplication / cilium assembly / ribosome-associated ubiquitin-dependent protein catabolic process / signal transduction in response to DNA damage / negative regulation of insulin receptor signaling pathway / Nuclear events stimulated by ALK signaling in cancer / protein K48-linked ubiquitination / regulation of cellular response to insulin stimulus / positive regulation of TORC1 signaling / transcription-coupled nucleotide-excision repair / post-translational protein modification / cellular response to amino acid stimulus / regulation of embryonic development / replication fork processing / negative regulation of canonical NF-kappaB signal transduction / regulation of mitotic cell cycle / site of DNA damage / Regulation of BACH1 activity / T cell activation / G1/S transition of mitotic cell cycle / negative regulation of canonical Wnt signaling pathway / epigenetic regulation of gene expression / Degradation of DVL / Degradation of CRY and PER proteins / nucleotide-excision repair / Degradation of GLI1 by the proteasome / regulation of autophagy / Recognition of DNA damage by PCNA-containing replication complex / microtubule cytoskeleton organization / GSK3B and BTRC:CUL1-mediated-degradation of NFE2L2 / Negative regulation of NOTCH4 signaling / RING-type E3 ubiquitin transferase / Hedgehog 'on' state / Vif-mediated degradation of APOBEC3G / FBXL7 down-regulates AURKA during mitotic entry and in early mitosis / Degradation of GLI2 by the proteasome / GLI3 is processed to GLI3R by the proteasome / GSK3B-mediated proteasomal degradation of PD-L1(CD274) / Evasion by RSV of host interferon responses / NOTCH1 Intracellular Domain Regulates Transcription / regulation of circadian rhythm / Degradation of beta-catenin by the destruction complex / cell population proliferation / Oxygen-dependent proline hydroxylation of Hypoxia-inducible Factor Alpha / DNA Damage Recognition in GG-NER / Constitutive Signaling by NOTCH1 PEST Domain Mutants / Constitutive Signaling by NOTCH1 HD+PEST Domain Mutants / Dual Incision in GG-NER / Transcription-Coupled Nucleotide Excision Repair (TC-NER) / Formation of TC-NER Pre-Incision Complex / SPOP-mediated proteasomal degradation of PD-L1(CD274)

Domain/homology:

: / : / : / : / : / : / CUL7/CUL9, N-terminal beta-barrel domain / CUL7/CUL9 ARM repeat domain / Cullin-9-like, TPR repeats / E3 ubiquitin-protein ligase ARIH2, N-terminal UBA domain / CPH domain / Mouse development and cellular proliferation protein Cullin-7 / CPH (CUL7, PARC, and HERC2) domain profile. / IBR domain, a half RING-finger domain / IBR domain / APC10/DOC domain / Anaphase-promoting complex, subunit 10 (APC10) / DOC domain profile. / Anaphase-promoting complex, subunit 10 (APC10) / IBR domain / In Between Ring fingers / TRIAD supradomain / TRIAD supradomain profile. / Zinc finger, RING-H2-type / RING-H2 zinc finger domain / : / Cullin, conserved site / Cullin family signature. / Cullin, N-terminal / Cullin protein, neddylation domain / Cullin / Cullin protein neddylation domain / Cullin alpha solenoid domain / : / Cullin alpha+beta domain / Cullin homology domain / Cullin homology domain superfamily / Cullin family profile. / Zinc finger, RING-type, conserved site / Zinc finger RING-type signature. / Galactose-binding-like domain superfamily / Zinc finger RING-type profile. / Zinc finger, RING-type / Armadillo-like helical / Armadillo-type fold / Zinc finger, RING/FYVE/PHD-type / Ribosomal protein L2, domain 2 / Winged helix-like DNA-binding domain superfamily

Component:

E3 ubiquitin-protein ligase RBX1 / Cullin-9

• Biological species: Homo sapiens (human)
• Method: single particle reconstruction / cryo EM / Resolution: 13.7 Å
• Authors: Hopf LVM / Horn-Ghetko D / Schulman BA

Funding support

European Union, Germany, 2 items

Organization	Grant number	Country
European Research Council (ERC)	789016	European Union
Max Planck Society		Germany

• Citation: Journal: Nat Struct Mol Biol / Year: 2024; Title: Noncanonical assembly, neddylation and chimeric cullin-RING/RBR ubiquitylation by the 1.8 MDa CUL9 E3 ligase complex.; Authors: Daniel Horn-Ghetko / Linus V M Hopf / Ishita Tripathi-Giesgen / Jiale Du / Sebastian Kostrhon / D Tung Vu / Viola Beier / Barbara Steigenberger / J Rajan Prabu / Luca Stier / Elias M Bruss / Matthias Mann / Yue Xiong / Brenda A Schulman / ; Abstract: Ubiquitin ligation is typically executed by hallmark E3 catalytic domains. Two such domains, 'cullin-RING' and 'RBR', are individually found in several hundred human E3 ligases, and collaborate with E2 enzymes to catalyze ubiquitylation. However, the vertebrate-specific CUL9 complex with RBX1 (also called ROC1), of interest due to its tumor suppressive interaction with TP53, uniquely encompasses both cullin-RING and RBR domains. Here, cryo-EM, biochemistry and cellular assays elucidate a 1.8-MDa hexameric human CUL9-RBX1 assembly. Within one dimeric subcomplex, an E2-bound RBR domain is activated by neddylation of its own cullin domain and positioning from the adjacent CUL9-RBX1 in trans. Our data show CUL9 as unique among RBX1-bound cullins in dependence on the metazoan-specific UBE2F neddylation enzyme, while the RBR domain protects it from deneddylation. Substrates are recruited to various upstream domains, while ubiquitylation relies on both CUL9's neddylated cullin and RBR domains achieving self-assembled and chimeric cullin-RING/RBR E3 ligase activity.

History

Deposition	Aug 16, 2023	-
Header (metadata) release	Apr 17, 2024	-
Map release	Apr 17, 2024	-
Update	Jul 31, 2024	-
Current status	Jul 31, 2024	Processing site: PDBe / Status: Released

Map

• File: Download / File: emd_18221.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
• Annotation: postprocess map
• Voxel size: X=Y=Z: 1.997 Å

Density

Contour Level	By AUTHOR: 0.022
Minimum - Maximum	-0.026007993 - 0.10900648
Average (Standard dev.)	0.0012027842 (±0.008378537)

• Symmetry: Space group: 1

Details

EMDB XML:

Map geometry	Axis order X Y Z Origin 0 0 0 Dimensions 256 256 256 Spacing 256 256 256
Cell	A=B=C: 511.232 Å α=β=γ: 90.0 °

Supplemental data

Additional map: refinement map

• File: emd_18221_additional_1.map
• Annotation: refinement map

Half map: #1

• File: emd_18221_half_map_1.map

Half map: #2

• File: emd_18221_half_map_2.map

Sample components

Entire : Structure of the hexameric CUL9-RBX1 complex with deletion of CUL...

• Entire: Name: Structure of the hexameric CUL9-RBX1 complex with deletion of CUL9 DOC domain

Components

• Complex: Structure of the hexameric CUL9-RBX1 complex with deletion of CUL9 DOC domain
  • Protein or peptide: Cullin-9
  • Protein or peptide: E3 ubiquitin-protein ligase RBX1

Supramolecule #1: Structure of the hexameric CUL9-RBX1 complex with deletion of CUL...

• Supramolecule: Name: Structure of the hexameric CUL9-RBX1 complex with deletion of CUL9 DOC domain; type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
• Source (natural): Organism: Homo sapiens (human)

Macromolecule #1: Cullin-9

• Macromolecule: Name: Cullin-9 / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
• Source (natural): Organism: Homo sapiens (human)
• Recombinant expression: Organism: Homo sapiens (human)

Sequence

String:

MVGERHAGDL MVPLGPRLQA YPEELIRQRP GHDGHPEYLI RWSVLKCGEV GKVGVEEGKA EHILMWLSAP EVYANCPGLL GERALSKGLQ HEPAGVSGSF PRDPGGLDEV AMGEMEADVQ ALVRRAARQL AESGTPSLTA AVLHTIHVLS AYASIGPLTG VFRETGALDL LMHMLCNPEP QIRRSAGKML QALAAHDAGS RAHVLLSLSQ QDGIEQHMDF DSRYTLLELF AETTSSEEHC MAFEGIHLPQ IPGKLLFSLV KRYLCVTSLL DQLNSSPELG AGDQSSPCAT REKSRGQREL EFSMAVGNLI SELVRSMGWA RNLSEQGMSP PRPTRSIFQP YISGPSLLLP TIVTTPRRQG WVFRQRSEFS SRSGYGEYVQ QTLQPGMRVR MLDDYEEISA GDEGEFRQSN NGIPPVQVFW QSTGRTYWVH WHMLEILGPE EATEDKASAA VEKGAGATVL GTAFPSWDWN PMDGLYPLPY LQPEPQKNER VGYLTQAEWW ELLFFIKKLD LCEQQPIFQN LWKNLDETLG EKALGEISVS VEMAESLLQV LSSRFEGSTL NDLLNSQIYT KYGLLSNEPS SSSTSRNHSC TPDPEEESKS EASFSEEETE SLKAKAEAPK TEAEPTKTRT ETPMAQSDSQ LFNQLLVTEG MTLPTEMKEA ASEMARALRG PGPRSSLDQH VAAVVATVQI SSLDTNLQLS GLSALSQAVE EVTERDHPLV RPDRSLREKL VKMLVELLTN QVGEKMVVVQ ALRLLYLLMT KHEWRPLFAR EGGIYAVLVC MQEYKTSVLV QQAGLAALKM LAVASSSEIP TFVTGRDSIH SLFDAQMTRE IFASIDSATR PGSESLLLTV PAAVILMLNT EGCSSAARNG LLLLNLLLCN HHTLGDQIIT QELRDTLFRH SGIAPRTEPM PTTRTILMML LNRYSEPPGS PERAALETPI IQGQDGSPEL LIRSLVGGPS AELLLDLERV LCREGSPGGA VRPLLKRLQQ ETQPFLLLLR TLDAPGPNKT LLLSVLRVIT RLLDFPEAMV LPWHEVLEPC LNCLSGPSSD SEIVQELTCF LHRLASMHKD YAVVLCCLGA KEILSKVLDK HSAQLLLGCE LRDLVTECEK YAQLYSNLTS SILAGCIQMV LGQIEDHRRT HQPINIPFFD VFLRHLCQGS SVEVKEGSGS GSGSGPKPTF WPLFREQLCR RTCLFYTIRA QAWSRDIAED HRRLLQLCPR LNRVLRHEQN FADRFLPDDE AAQALGKTCW EALVSPLVQN ITSPDAEGVS ALGWLLDQYL EQRETSRNPL SRAASFASRV RRLCHLLVHV EPPPGPSPEP STRPFSKNSK GRDRSPAPSP VLPSSSLRNI TQCWLSVVQE QVSRFLAAAW RAPDFVPRYC KLYEHLQRAG SELFGPRAAF MLALRSGFSG ALLQQSFLTA AHMSEQFARY IDQQIQGGLI GGAPGVEMLG QLQRHLEPIM VLSGLELATT FEHFYQHYMA DRLLSFGSSW LEGAVLEQIG LCFPNRLPQL MLQSLSTSEE LQRQFHLFQL QRLDKLFLEQ EDEEEKRLEE EEEEEEEEEA EKELFIEDPS PAISILVLSP RCWPVSPLCY LYHPRKCLPT EFCDALDRFS SFYSQSQNHP VLDMGPHRRL QWTWLGRAEL QFGKQILHVS TVQMWLLLKF NQTEEVSVET LLKDSDLSPE LLLQALVPLT SGNGPLTLHE GQDFPHGGVL RLHEPGPQRS GEALWLIPPQ AYLNVEKDEG RTLEQKRNLL SCLLVRILKA HGEKGLHIDQ LVCLVLEAWQ KGPNPPGTLG HTVAGGVACT STDVLSCILH LLGQGYVKRR DDRPQILMYA APEPMGPCRG QADVPFCGSQ SETSKPSPEA VATLASLQLP AGRTMSPQEV EGLMKQTVRQ VQETLNLEPD VAQHLLAHSH WGAEQLLQSY SEDPEPLLLA AGLCVHQAQA VPVRPDHCPV CVSPLGCDDD LPSLCCMHYC CKSCWNEYLT TRIEQNLVLN CTCPIADCPA QPTGAFIRAI VSSPEVISKY EKALLRGYVE SCSNLTWCTN PQGCDRILCR QGLGCGTTCS KCGWASCFNC SFPEAHYPAS CGHMSQWVDD GGYYDGMSVE AQSKHLAKLI SKRCPSCQAP IEKNEGCLHM TCAKCNHGFC WRCLKSWKPN HKDYYNCSAM VSKAARQEKR FQDYNERCTF HHQAREFAVN LRNRVSAIHE VPPPRSFTFL NDACQGLEQA RKVLAYACVY SFYSQDAEYM DVVEQQTENL ELHTNALQIL LEETLLRCRD LASSLRLLRA DCLSTGMELL RRIQERLLAI LQHSAQDFRV GLQSPSVEAW EAKGPNMPGS QPQASSGPEA EEEEEDDEDD VPEWQQDEFD EELDNDSFSY DESENLDQET FFFGDEEEDE DEAYD

UniProtKB: Cullin-9

Macromolecule #2: E3 ubiquitin-protein ligase RBX1

• Macromolecule: Name: E3 ubiquitin-protein ligase RBX1 / type: protein_or_peptide / ID: 2 / Enantiomer: LEVO
• Source (natural): Organism: Homo sapiens (human)
• Recombinant expression: Organism: Homo (humans)

Sequence

String:

MDVDTPSGTN SGAGKKRFEV KKWNAVALWA WDIVVDNCAI CRNHIMDLCI ECQANQ ASA TSEECTVAWG VCNHAFHFHC ISRWLKTRQV CPLDNREWEF QKYGH

UniProtKB: E3 ubiquitin-protein ligase RBX1

Experimental details

Structure determination

• Method: cryo EM
• Processing: single particle reconstruction
• Aggregation state: particle

Sample preparation

• Concentration: 3 mg/mL
• Buffer: pH: 7.5
• Vitrification: Cryogen name: ETHANE

Electron microscopy

• Microscope: FEI TALOS ARCTICA
• Image recording: Film or detector model: FEI FALCON III (4k x 4k) / Average electron dose: 60.0 e/Ų
• Electron beam: Acceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
• Electron optics: Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 3.3000000000000003 µm / Nominal defocus min: 1.2 µm
• Experimental equipment: Model: Talos Arctica / Image courtesy: FEI Company

Image processing

• Startup model: Type of model: NONE
• Final reconstruction: Resolution.type: BY AUTHOR / Resolution: 13.7 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 29312
• Initial angle assignment: Type: MAXIMUM LIKELIHOOD
• Final angle assignment: Type: MAXIMUM LIKELIHOOD