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- PDB-9b62: Human RANBP2/RAN(GTP)/RANGAP1-SUMO1/UBC9/CRM1/RAN(GTP) - composit... -

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Basic information

Entry
Database: PDB / ID: 9b62
TitleHuman RANBP2/RAN(GTP)/RANGAP1-SUMO1/UBC9/CRM1/RAN(GTP) - composite map and model
Components
  • E3 SUMO-protein ligase RanBP2
  • Exportin-1
  • GTP-binding nuclear protein Ran
  • Ran GTPase-activating protein 1
  • SUMO-conjugating enzyme UBC9
  • Small ubiquitin-related modifier 1
KeywordsTRANSPORT PROTEIN / karyopherin / SUMO E3 / SUMO E2 / transporter / GTPase / GTPase activating protein / exportin / G-protein
Function / homology
Function and homology information


: / cellular response to vasopressin / SUMO conjugating enzyme activity / cytoplasmic periphery of the nuclear pore complex / RING-like zinc finger domain binding / HuR (ELAVL1) binds and stabilizes mRNA / SUMO ligase complex / SUMO ligase activity / protein localization to nuclear pore / negative regulation of transcription by transcription factor localization ...: / cellular response to vasopressin / SUMO conjugating enzyme activity / cytoplasmic periphery of the nuclear pore complex / RING-like zinc finger domain binding / HuR (ELAVL1) binds and stabilizes mRNA / SUMO ligase complex / SUMO ligase activity / protein localization to nuclear pore / negative regulation of transcription by transcription factor localization / transferase complex / annulate lamellae / SUMOylation of nuclear envelope proteins / SUMO is proteolytically processed / Negative regulation of activity of TFAP2 (AP-2) family transcription factors / negative regulation of delayed rectifier potassium channel activity / SUMO is conjugated to E1 (UBA2:SAE1) / regulation of proteasomal ubiquitin-dependent protein catabolic process / negative regulation of potassium ion transmembrane transporter activity / PML body organization / SUMO is transferred from E1 to E2 (UBE2I, UBC9) / nuclear stress granule / Vitamin D (calciferol) metabolism / pre-miRNA export from nucleus / RNA nuclear export complex / nuclear pore cytoplasmic filaments / mitotic nuclear membrane reassembly / snRNA import into nucleus / synaptonemal complex / regulation of centrosome duplication / nuclear export signal receptor activity / Nuclear Pore Complex (NPC) Disassembly / small protein activating enzyme binding / negative regulation of action potential / nuclear inclusion body / Transport of Ribonucleoproteins into the Host Nucleus / nuclear pore nuclear basket / Regulation of Glucokinase by Glucokinase Regulatory Protein / Defective TPR may confer susceptibility towards thyroid papillary carcinoma (TPC) / Regulation of cholesterol biosynthesis by SREBP (SREBF) / SUMOylation of DNA methylation proteins / Transport of the SLBP independent Mature mRNA / Transport of the SLBP Dependant Mature mRNA / NS1 Mediated Effects on Host Pathways / SUMOylation of immune response proteins / regulation of calcium ion transmembrane transport / XY body / SUMOylation of SUMOylation proteins / NLS-bearing protein import into nucleus / regulation of protein export from nucleus / activation of GTPase activity / Transport of Mature mRNA Derived from an Intronless Transcript / protein localization to nucleolus / Rev-mediated nuclear export of HIV RNA / Maturation of nucleoprotein / negative regulation of protein export from nucleus / Nuclear import of Rev protein / SUMOylation of RNA binding proteins / nuclear export / regulation of cardiac muscle cell contraction / NEP/NS2 Interacts with the Cellular Export Machinery / kinase activator activity / Transport of Mature mRNA derived from an Intron-Containing Transcript / Transferases; Acyltransferases; Aminoacyltransferases / GTP metabolic process / tRNA processing in the nucleus / SUMO transferase activity / Postmitotic nuclear pore complex (NPC) reformation / negative regulation of DNA binding / nucleocytoplasmic transport / aggresome / Maturation of nucleoprotein / centrosome localization / MicroRNA (miRNA) biogenesis / Viral Messenger RNA Synthesis / regulation of gluconeogenesis / DNA metabolic process / negative regulation of protein import into nucleus / SUMOylation of ubiquitinylation proteins / cellular response to cadmium ion / ubiquitin-specific protease binding / Vpr-mediated nuclear import of PICs / transcription factor binding / Maturation of hRSV A proteins / roof of mouth development / ubiquitin-like protein ligase binding / SUMOylation of transcription factors / mitotic sister chromatid segregation / protein sumoylation / SUMOylation of DNA replication proteins / postsynaptic cytosol / ribosomal large subunit export from nucleus / Estrogen-dependent nuclear events downstream of ESR-membrane signaling / protein localization to nucleus / Regulation of HSF1-mediated heat shock response / response to axon injury / mRNA transport / viral process / potassium channel regulator activity / Regulation of IFNG signaling
Similarity search - Function
Ran-GTPase activating protein 1, C-terminal / Ran-GTPase activating protein 1, C-terminal domain superfamily / RanGAP1 C-terminal domain / : / Nup358/RanBP2 E3 ligase domain / Nup358/RanBP2 E3 ligase domain / Small ubiquitin-related modifier 1, Ubl domain / Exportin-1, repeat 3 / Chromosome region maintenance repeat / Exportin-1, repeat 2 ...Ran-GTPase activating protein 1, C-terminal / Ran-GTPase activating protein 1, C-terminal domain superfamily / RanGAP1 C-terminal domain / : / Nup358/RanBP2 E3 ligase domain / Nup358/RanBP2 E3 ligase domain / Small ubiquitin-related modifier 1, Ubl domain / Exportin-1, repeat 3 / Chromosome region maintenance repeat / Exportin-1, repeat 2 / Chromosome region maintenance or exportin repeat / CRM1 / Exportin repeat 2 / CRM1 / Exportin repeat 3 / CRM1 C terminal / Exportin-1, C-terminal / CRM1 C terminal / Exportin-1/5 / Exportin-1/Importin-beta-like / Exportin 1-like protein / Ran binding protein RanBP1-like / Ran binding domain / RanBP1 domain / Ran binding domain type 1 profile. / Ran-binding domain / small GTPase Ran family profile. / Ran GTPase / Importin-beta N-terminal domain / Importin-beta N-terminal domain / Importin-beta N-terminal domain profile. / Importin-beta, N-terminal domain / Rad60/SUMO-like domain / Ubiquitin-2 like Rad60 SUMO-like / Leucine rich repeat, ribonuclease inhibitor type / : / Zinc finger domain / Leucine Rich repeat / Zn-finger in Ran binding protein and others / Zinc finger RanBP2 type profile. / Zinc finger, RanBP2-type superfamily / Zinc finger RanBP2-type signature. / Zinc finger, RanBP2-type / Ubiquitin-conjugating enzyme, active site / Ubiquitin-conjugating (UBC) active site signature. / Cyclophilin-type peptidyl-prolyl cis-trans isomerase, conserved site / Cyclophilin-type peptidyl-prolyl cis-trans isomerase signature. / Ubiquitin-conjugating enzyme E2 / Ubiquitin-conjugating enzyme / Ubiquitin-conjugating (UBC) core domain profile. / Ubiquitin-conjugating enzyme E2, catalytic domain homologues / Cyclophilin-type peptidyl-prolyl cis-trans isomerase domain profile. / Cyclophilin-type peptidyl-prolyl cis-trans isomerase domain / Cyclophilin type peptidyl-prolyl cis-trans isomerase/CLD / Cyclophilin-like domain superfamily / Ubiquitin-conjugating enzyme/RWD-like / Ran (Ras-related nuclear proteins) /TC4 subfamily of small GTPases / TPR repeat region circular profile. / TPR repeat profile. / Tetratricopeptide repeats / Tetratricopeptide repeat / Leucine-rich repeat / Rho (Ras homology) subfamily of Ras-like small GTPases / Ras subfamily of RAS small GTPases / Small GTPase / Ras family / Rab subfamily of small GTPases / Leucine-rich repeat domain superfamily / Armadillo-like helical / Small GTP-binding protein domain / Ubiquitin homologues / Tetratricopeptide-like helical domain superfamily / Ubiquitin domain profile. / Ubiquitin-like domain / PH-like domain superfamily / Armadillo-type fold / Ubiquitin-like domain superfamily / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
GUANOSINE-5'-TRIPHOSPHATE / Exportin-1 / Ran GTPase-activating protein 1 / E3 SUMO-protein ligase RanBP2 / GTP-binding nuclear protein Ran / Small ubiquitin-related modifier 1 / SUMO-conjugating enzyme UBC9
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.9 Å
AuthorsLima, C.D. / DiMattia, M.A.
Funding support United States, 2items
OrganizationGrant numberCountry
Howard Hughes Medical Institute (HHMI) United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM118080 United States
CitationJournal: bioRxiv / Year: 2024
Title: Structural basis for a nucleoporin exportin complex between RanBP2, SUMO1-RanGAP1, the E2 Ubc9, Crm1 and the Ran GTPase.
Authors: Vladimir Baytshtok / Michael A DiMattia / Christopher D Lima /
Abstract: The human nucleoporin RanBP2/Nup358 interacts with SUMO1-modified RanGAP1 and the SUMO E2 Ubc9 at the nuclear pore complex (NPC) to promote export and disassembly of exportin Crm1/Ran(GTP)/cargo ...The human nucleoporin RanBP2/Nup358 interacts with SUMO1-modified RanGAP1 and the SUMO E2 Ubc9 at the nuclear pore complex (NPC) to promote export and disassembly of exportin Crm1/Ran(GTP)/cargo complexes. In mitosis, RanBP2/SUMO1-RanGAP1/Ubc9 remains intact after NPC disassembly and is recruited to kinetochores and mitotic spindles by Crm1 where it contributes to mitotic progression. Interestingly, RanBP2 binds SUMO1-RanGAP1/Ubc9 via motifs that also catalyze SUMO E3 ligase activity. Here, we resolve cryo-EM structures of a RanBP2 C-terminal fragment in complex with Crm1, SUMO1-RanGAP1/Ubc9, and two molecules of Ran(GTP). These structures reveal several unanticipated interactions with Crm1 including a nuclear export signal (NES) for RanGAP1, the deletion of which mislocalizes RanGAP1 and the Ran GTPase in cells. Our structural and biochemical results support models in which RanBP2 E3 ligase activity is dependent on Crm1, the RanGAP1 NES and Ran GTPase cycling.
History
DepositionMar 23, 2024Deposition site: RCSB / Processing site: RCSB
Revision 1.0Oct 16, 2024Provider: repository / Type: Initial release
Revision 1.1Apr 30, 2025Group: Data collection / Database references / Category: citation / citation_author / em_admin
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _citation_author.identifier_ORCID / _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Exportin-1
B: GTP-binding nuclear protein Ran
C: SUMO-conjugating enzyme UBC9
D: Small ubiquitin-related modifier 1
E: E3 SUMO-protein ligase RanBP2
F: GTP-binding nuclear protein Ran
G: Ran GTPase-activating protein 1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)337,26211
Polymers336,1677
Non-polymers1,0954
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration, co-migrates as a 1:1:1:1:1:2 complex between Nup358/RanBP2:RanGAP1-SUMO1:UBC9:CRM1:RAN, cross-linking, Substochimetric crosslinking used. While cross-linking can occur ...Evidence: gel filtration, co-migrates as a 1:1:1:1:1:2 complex between Nup358/RanBP2:RanGAP1-SUMO1:UBC9:CRM1:RAN, cross-linking, Substochimetric crosslinking used. While cross-linking can occur between all of the subunits, SDS-PAGE analysis of the crosslinked material suggests substochiometric crosslinking with most of the material not shifted or crosslinked, electron microscopy, Structure determined by single particle cryo-EM revealed the expected assembly (and subcomplexes)
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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Protein , 6 types, 7 molecules ABFCDEG

#1: Protein Exportin-1 / Exp1 / Chromosome region maintenance 1 protein homolog


Mass: 123874.781 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Details: SHM N-terminal amino acids remnants of expression tag
Source: (gene. exp.) Homo sapiens (human) / Gene: XPO1, CRM1 / Production host: Escherichia coli (E. coli) / References: UniProt: O14980
#2: Protein GTP-binding nuclear protein Ran / Androgen receptor-associated protein 24 / GTPase Ran / Ras-like protein TC4 / Ras-related nuclear protein


Mass: 25012.764 Da / Num. of mol.: 2 / Mutation: Q69L
Source method: isolated from a genetically manipulated source
Details: GSHMAS N-terminal amino acids residual after removal of purification epitope tag
Source: (gene. exp.) Homo sapiens (human) / Gene: RAN, ARA24, OK/SW-cl.81 / Production host: Escherichia coli (E. coli)
References: UniProt: P62826, Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement
#3: Protein SUMO-conjugating enzyme UBC9 / RING-type E3 SUMO transferase UBC9 / SUMO-protein ligase / Ubiquitin carrier protein 9 / Ubiquitin ...RING-type E3 SUMO transferase UBC9 / SUMO-protein ligase / Ubiquitin carrier protein 9 / Ubiquitin carrier protein I / Ubiquitin-conjugating enzyme E2 I / Ubiquitin-protein ligase I / p18


Mass: 18313.092 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Details: GSH N-terminal amino acids left over after cleavage of affinity tag
Source: (gene. exp.) Homo sapiens (human) / Gene: UBE2I, UBC9, UBCE9 / Production host: Escherichia coli (E. coli)
References: UniProt: P63279, Transferases; Acyltransferases; Aminoacyltransferases
#4: Protein Small ubiquitin-related modifier 1 / SUMO-1 / GAP-modifying protein 1 / GMP1 / SMT3 homolog 3 / Sentrin / Ubiquitin-homology domain ...SUMO-1 / GAP-modifying protein 1 / GMP1 / SMT3 homolog 3 / Sentrin / Ubiquitin-homology domain protein PIC1 / Ubiquitin-like protein SMT3C / Smt3C / Ubiquitin-like protein UBL1


Mass: 11532.003 Da / Num. of mol.: 1 / Mutation: Q94P
Source method: isolated from a genetically manipulated source
Details: GSHM N-terminal amino acids left over after cleavage of affinity tag
Source: (gene. exp.) Homo sapiens (human) / Gene: SUMO1, SMT3C, SMT3H3, UBL1, OK/SW-cl.43 / Production host: Escherichia coli (E. coli) / References: UniProt: P63165
#5: Protein E3 SUMO-protein ligase RanBP2 / 358 kDa nucleoporin / Nuclear pore complex protein Nup358 / Nucleoporin Nup358 / Ran-binding ...358 kDa nucleoporin / Nuclear pore complex protein Nup358 / Nucleoporin Nup358 / Ran-binding protein 2 / RanBP2 / p270


Mass: 68399.422 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Details: GSH N-terminal amino acids left after cleavage of affinity tag The residues that are misaligned in your alignment should be aligned to residue 66 with a gap after. please fix
Source: (gene. exp.) Homo sapiens (human) / Gene: RANBP2, NUP358 / Production host: Escherichia coli (E. coli)
References: UniProt: P49792, Transferases; Acyltransferases; Aminoacyltransferases
#6: Protein Ran GTPase-activating protein 1 / RanGAP1


Mass: 64022.172 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Details: GSHM N-terminal amino acids left after cleavage of affinity tag
Source: (gene. exp.) Homo sapiens (human) / Gene: RANGAP1, KIAA1835, SD / Production host: Escherichia coli (E. coli) / References: UniProt: P46060

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Non-polymers , 2 types, 4 molecules

#7: Chemical ChemComp-GTP / GUANOSINE-5'-TRIPHOSPHATE


Mass: 523.180 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C10H16N5O14P3 / Comment: GTP, energy-carrying molecule*YM
#8: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Mg

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Details

Has ligand of interestN
Has protein modificationN

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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

ComponentName: Human RANBP2/RAN(GTP)/RANGAP1-SUMO1/UBC9/CRM1/RAN(GTP) complex
Type: COMPLEX
Details: SUMO1 Gly97 is covalently linked to RANGAP1 Lys524 through a isopeptide bond
Entity ID: #1-#6 / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 8
Details: 20 mM Tris-Cl pH 8.0, 50 mM NaCl, 0.1 mM TCEP supplemented with 0.02% (v/v) IGEPAL CA-630
SpecimenConc.: 8 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES / Details: Gel filtered - sample was monodisperse
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 295 K / Details: 30 s wait time, blot for 2.5 s before plunging

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 3000 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 10 sec. / Electron dose: 85.2 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K2 SUMMIT (4k x 4k)

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Processing

EM software
IDNameVersionCategory
1RELIONparticle selection
2SerialEMimage acquisition
4GctfCTF correction
9PHENIX1.19.2_4158:model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 895509
Details: This is a composite map so numbers of particles per map component differs - after selection of particles containing Crm1 there were 704835 unique particles - after selection of particles ...Details: This is a composite map so numbers of particles per map component differs - after selection of particles containing Crm1 there were 704835 unique particles - after selection of particles containing Crm1-Ran(GTP) there were 534708 unique particles
3D reconstructionResolution: 2.9 Å / Resolution method: OTHER / Num. of particles: 534708
Details: Composite map - estimated best average resolution of FR maps 2.9 Related particles and maps used to generate the composite listed in Related Entries
Symmetry type: POINT
Atomic model buildingProtocol: OTHER / Space: REAL
Details: Atomic model was manually fit and refined using composite map, and manually inspected using Coot
Atomic model building
IDPDB-ID 3D fitting-IDAccession codeInitial refinement model-IDSource nameType
13uin

3uin

13uin1PDBexperimental model
23gjx

3gjx

13gjx2PDBexperimental model
34l6e

4l6e

14l6e3PDBexperimental model
41k5d

1k5d

11k5d4PDBexperimental model
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00319677
ELECTRON MICROSCOPYf_angle_d0.43626627
ELECTRON MICROSCOPYf_dihedral_angle_d10.5897337
ELECTRON MICROSCOPYf_chiral_restr0.0373006
ELECTRON MICROSCOPYf_plane_restr0.0033405

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