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- PDB-6v4x: Cryo-EM structure of an active human histone pre-mRNA 3'-end proc... -

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Entry
Database: PDB / ID: 6v4x
TitleCryo-EM structure of an active human histone pre-mRNA 3'-end processing machinery at 3.2 Angstrom resolution
Components
  • (Cleavage and polyadenylation specificity factor subunit ...) x 2
  • (Small nuclear ribonucleoprotein ...) x 4
  • (U7 snRNA-associated Sm-like protein ...) x 2
  • Small nuclear ribonucleoprotein-associated proteins B and B'
  • Symplekin
  • U7 snRNA
  • modified H2a pre-mRNA
KeywordsRNA BINDING PROTEIN/RNA / endonuclease / active CPSF73 / U7 snRNP / 3'-end processing / RIBONUCLEASE / RNA BINDING PROTEIN-RNA complex
Function / homology
Function and homology information


cytoplasmic U snRNP body / mRNA 3'-end processing by stem-loop binding and cleavage / co-transcriptional mRNA 3'-end processing, cleavage and polyadenylation pathway / Processing of Intronless Pre-mRNAs / 5'-3' RNA exonuclease activity / mRNA cleavage and polyadenylation specificity factor complex / U2 snRNP binding / nuclear stress granule / regulation of chromatin organization / U7 snRNA binding ...cytoplasmic U snRNP body / mRNA 3'-end processing by stem-loop binding and cleavage / co-transcriptional mRNA 3'-end processing, cleavage and polyadenylation pathway / Processing of Intronless Pre-mRNAs / 5'-3' RNA exonuclease activity / mRNA cleavage and polyadenylation specificity factor complex / U2 snRNP binding / nuclear stress granule / regulation of chromatin organization / U7 snRNA binding / histone pre-mRNA DCP binding / U7 snRNP / histone pre-mRNA 3'end processing complex / SLBP independent Processing of Histone Pre-mRNAs / SLBP Dependent Processing of Replication-Dependent Histone Pre-mRNAs / protein methylation / U12-type spliceosomal complex / 7-methylguanosine cap hypermethylation / U1 snRNP binding / Hydrolases; Acting on ester bonds; Endoribonucleases producing 3'-phosphomonoesters / methylosome / mRNA 3'-end processing / pICln-Sm protein complex / Transport of Mature mRNA Derived from an Intronless Transcript / snRNP binding / SMN-Sm protein complex / small nuclear ribonucleoprotein complex / mRNA 3'-end processing / spliceosomal tri-snRNP complex / P granule / telomerase holoenzyme complex / U2-type precatalytic spliceosome / commitment complex / telomerase RNA binding / U2-type spliceosomal complex / U2-type prespliceosome assembly / U2-type catalytic step 2 spliceosome / RNA Polymerase II Transcription Termination / U2 snRNP / U1 snRNP / U4 snRNP / U2-type prespliceosome / precatalytic spliceosome / spliceosomal complex assembly / mRNA Splicing - Minor Pathway / Processing of Capped Intron-Containing Pre-mRNA / U5 snRNP / bicellular tight junction / positive regulation of G1/S transition of mitotic cell cycle / spliceosomal snRNP assembly / Cajal body / negative regulation of protein binding / U4/U6 x U5 tri-snRNP complex / catalytic step 2 spliceosome / RNA endonuclease activity / mRNA Splicing - Major Pathway / RNA splicing / spliceosomal complex / mRNA splicing, via spliceosome / mRNA processing / snRNP Assembly / SARS-CoV-2 modulates host translation machinery / cytoskeleton / postsynapse / cell adhesion / nuclear body / ribonucleoprotein complex / glutamatergic synapse / enzyme binding / RNA binding / nucleoplasm / metal ion binding / nucleus / membrane / plasma membrane / cytoplasm / cytosol
Similarity search - Function
: / Sm-like protein Lsm11, middle domain / U7 snRNA-associated Sm-like protein Lsm11 / Cleavage and polyadenylation specificity factor 2, C-terminal / CPSF2, metallo-hydrolase domain / Cleavage and polyadenylation factor 2 C-terminal / Cleavage and polyadenylation specificity factor subunit 2 / Pre-mRNA 3'-end-processing endonuclease polyadenylation factor C-term / Pre-mRNA 3'-end-processing endonuclease polyadenylation factor C-term / CPSF73-100_C ...: / Sm-like protein Lsm11, middle domain / U7 snRNA-associated Sm-like protein Lsm11 / Cleavage and polyadenylation specificity factor 2, C-terminal / CPSF2, metallo-hydrolase domain / Cleavage and polyadenylation factor 2 C-terminal / Cleavage and polyadenylation specificity factor subunit 2 / Pre-mRNA 3'-end-processing endonuclease polyadenylation factor C-term / Pre-mRNA 3'-end-processing endonuclease polyadenylation factor C-term / CPSF73-100_C / Symplekin/Pta1 / Symplekin C-terminal / Symplekin/Pta1, N-terminal / Symplekin/PTA1 N-terminal / Symplekin tight junction protein C terminal / Rossmann fold - #10890 / Metallo-beta-lactamase superfamily domain / : / Beta-Casp domain / Beta-Casp domain / Beta-Casp domain / Zn-dependent metallo-hydrolase, RNA specificity domain / Zn-dependent metallo-hydrolase RNA specificity domain / Small ribonucleoprotein associated, SmB/SmN / SH3 type barrels. - #100 / Small nuclear ribonucleoprotein Sm D3 / Small nuclear ribonucleoprotein E / Small nuclear ribonucleoprotein G / Small nuclear ribonucleoprotein F / Sm-like protein Lsm7/SmG / Like-Sm (LSM) domain containing protein, LSm4/SmD1/SmD3 / Sm-like protein Lsm6/SmF / LSM domain / LSM domain, eukaryotic/archaea-type / snRNP Sm proteins / : / Sm domain profile. / LSM domain superfamily / Leucine-rich Repeat Variant / Leucine-rich Repeat Variant / Metallo-beta-lactamase superfamily / Metallo-beta-lactamase superfamily / Metallo-beta-lactamase / Ribonuclease Z/Hydroxyacylglutathione hydrolase-like / SH3 type barrels. / Armadillo-like helical / Alpha Horseshoe / Armadillo-type fold / Roll / Rossmann fold / 3-Layer(aba) Sandwich / Mainly Beta / Mainly Alpha / Alpha Beta
Similarity search - Domain/homology
: / RNA / RNA (> 10) / Small nuclear ribonucleoprotein-associated proteins B and B' / Small nuclear ribonucleoprotein E / Small nuclear ribonucleoprotein F / Small nuclear ribonucleoprotein G / Small nuclear ribonucleoprotein Sm D3 / U7 snRNA-associated Sm-like protein LSm11 / Symplekin ...: / RNA / RNA (> 10) / Small nuclear ribonucleoprotein-associated proteins B and B' / Small nuclear ribonucleoprotein E / Small nuclear ribonucleoprotein F / Small nuclear ribonucleoprotein G / Small nuclear ribonucleoprotein Sm D3 / U7 snRNA-associated Sm-like protein LSm11 / Symplekin / U7 snRNA-associated Sm-like protein LSm10 / Cleavage and polyadenylation specificity factor subunit 2 / Cleavage and polyadenylation specificity factor subunit 3
Similarity search - Component
Biological speciesHomo sapiens (human)
Mus musculus (house mouse)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.2 Å
AuthorsSun, Y. / Zhang, Y. / Walz, T. / Tong, L.
Funding support United States, 2items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM118093 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01GM029832 United States
CitationJournal: Science / Year: 2020
Title: Structure of an active human histone pre-mRNA 3'-end processing machinery.
Authors: Yadong Sun / Yixiao Zhang / Wei Shen Aik / Xiao-Cui Yang / William F Marzluff / Thomas Walz / Zbigniew Dominski / Liang Tong /
Abstract: The 3'-end processing machinery for metazoan replication-dependent histone precursor messenger RNAs (pre-mRNAs) contains the U7 small nuclear ribonucleoprotein and shares the key cleavage module with ...The 3'-end processing machinery for metazoan replication-dependent histone precursor messenger RNAs (pre-mRNAs) contains the U7 small nuclear ribonucleoprotein and shares the key cleavage module with the canonical cleavage and polyadenylation machinery. We reconstituted an active human histone pre-mRNA processing machinery using 13 recombinant proteins and two RNAs and determined its structure by cryo-electron microscopy. The overall structure is highly asymmetrical and resembles an amphora with one long handle. We captured the pre-mRNA in the active site of the endonuclease, the 73-kilodalton subunit of the cleavage and polyadenylation specificity factor, poised for cleavage. The endonuclease and the entire cleavage module undergo extensive rearrangements for activation, triggered through the recognition of the duplex between the authentic pre-mRNA and U7 small nuclear RNA (snRNA). Our study also has notable implications for understanding canonical and snRNA 3'-end processing.
History
DepositionDec 2, 2019Deposition site: RCSB / Processing site: RCSB
Revision 1.0Feb 19, 2020Provider: repository / Type: Initial release
Revision 1.0Feb 19, 2020Data content type: EM metadata / Data content type: EM metadata / Provider: repository / Type: Initial release
Revision 1.0Feb 19, 2020Data content type: Image / Data content type: Image / Provider: repository / Type: Initial release
Revision 1.0Feb 19, 2020Data content type: Primary map / Data content type: Primary map / Provider: repository / Type: Initial release
Revision 1.0Feb 19, 2020Data content type: EM metadata / Data content type: EM metadata / Provider: repository / Type: Initial release
Revision 1.0Feb 19, 2020Data content type: Image / Data content type: Image / Provider: repository / Type: Initial release
Revision 1.0Feb 19, 2020Data content type: Primary map / Data content type: Primary map / Provider: repository / Type: Initial release
Revision 1.0Feb 19, 2020Data content type: EM metadata / Data content type: EM metadata / Provider: repository / Type: Initial release
Revision 1.0Feb 19, 2020Data content type: Image / Data content type: Image / Provider: repository / Type: Initial release
Revision 1.0Feb 19, 2020Data content type: Primary map / Data content type: Primary map / Provider: repository / Type: Initial release
Revision 1.1Mar 16, 2022Group: Author supporting evidence / Database references / Derived calculations
Category: database_2 / pdbx_audit_support / struct_conn
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _pdbx_audit_support.funding_organization / _struct_conn.ptnr1_auth_asym_id / _struct_conn.ptnr1_auth_comp_id / _struct_conn.ptnr1_auth_seq_id / _struct_conn.ptnr1_label_asym_id / _struct_conn.ptnr1_label_atom_id / _struct_conn.ptnr1_label_comp_id / _struct_conn.ptnr1_label_seq_id / _struct_conn.ptnr2_auth_asym_id / _struct_conn.ptnr2_auth_comp_id / _struct_conn.ptnr2_auth_seq_id / _struct_conn.ptnr2_label_asym_id / _struct_conn.ptnr2_label_atom_id / _struct_conn.ptnr2_label_comp_id / _struct_conn.ptnr2_label_seq_id
Revision 1.2May 29, 2024Group: Data collection / Category: chem_comp_atom / chem_comp_bond
Revision 1.3Jun 4, 2025Group: Data collection / Structure summary / Category: em_admin / em_software / pdbx_entry_details
Item: _em_admin.last_update / _em_software.name / _pdbx_entry_details.has_protein_modification
Revision 1.1Jun 4, 2025Data content type: EM metadata / Data content type: EM metadata / EM metadata / Group: Data processing / Experimental summary / Data content type: EM metadata / EM metadata / Category: em_admin / em_software / Data content type: EM metadata / EM metadata / Item: _em_admin.last_update / _em_software.name

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Structure visualization

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Assembly

Deposited unit
A: Small nuclear ribonucleoprotein Sm D3
B: Small nuclear ribonucleoprotein-associated proteins B and B'
F: Small nuclear ribonucleoprotein F
E: Small nuclear ribonucleoprotein E
G: Small nuclear ribonucleoprotein G
C: U7 snRNA-associated Sm-like protein LSm10
D: U7 snRNA-associated Sm-like protein LSm11
H: Cleavage and polyadenylation specificity factor subunit 3
I: Cleavage and polyadenylation specificity factor subunit 2
J: Symplekin
Z: U7 snRNA
Y: modified H2a pre-mRNA
hetero molecules


Theoretical massNumber of molelcules
Total (without water)422,25514
Polymers422,12412
Non-polymers1312
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area38450 Å2
ΔGint-248 kcal/mol
Surface area78240 Å2

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Components

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Small nuclear ribonucleoprotein ... , 4 types, 4 molecules AFEG

#1: Protein Small nuclear ribonucleoprotein Sm D3 / Sm-D3 / snRNP core protein D3


Mass: 16111.671 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: SNRPD3 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P62318
#3: Protein Small nuclear ribonucleoprotein F / snRNP-F / Sm protein F / SmF


Mass: 9734.171 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: SNRPF, PBSCF / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P62306
#4: Protein Small nuclear ribonucleoprotein E / snRNP-E / Sm protein E / SmE


Mass: 10817.601 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: SNRPE / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P62304
#5: Protein Small nuclear ribonucleoprotein G / snRNP-G / Sm protein G / SmG


Mass: 9579.236 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: SNRPG, PBSCG / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P62308

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Protein , 2 types, 2 molecules BJ

#2: Protein Small nuclear ribonucleoprotein-associated proteins B and B' / snRNP-B / Sm protein B/B' / SmB/B'


Mass: 10911.931 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: SNRPB, COD, SNRPB1 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P14678
#10: Protein Symplekin


Mass: 120355.125 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: SYMPK, SPK / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q92797

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U7 snRNA-associated Sm-like protein ... , 2 types, 2 molecules CD

#6: Protein U7 snRNA-associated Sm-like protein LSm10


Mass: 14102.057 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: LSM10 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q969L4
#7: Protein U7 snRNA-associated Sm-like protein LSm11


Mass: 28609.793 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: LSM11 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P83369

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Cleavage and polyadenylation specificity factor subunit ... , 2 types, 2 molecules HI

#8: Protein Cleavage and polyadenylation specificity factor subunit 3 / Cleavage and polyadenylation specificity factor 73 kDa subunit / CPSF 73 kDa subunit / mRNA 3'-end- ...Cleavage and polyadenylation specificity factor 73 kDa subunit / CPSF 73 kDa subunit / mRNA 3'-end-processing endonuclease CPSF-73


Mass: 77580.883 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CPSF3, CPSF73 / Production host: Trichoplusia ni (cabbage looper)
References: UniProt: Q9UKF6, Hydrolases; Acting on ester bonds; Endoribonucleases producing 3'-phosphomonoesters
#9: Protein Cleavage and polyadenylation specificity factor subunit 2 / Cleavage and polyadenylation specificity factor 100 kDa subunit / CPSF 100 kDa subunit


Mass: 88597.734 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CPSF2, CPSF100, KIAA1367 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9P2I0

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RNA chain , 2 types, 2 molecules ZY

#11: RNA chain U7 snRNA


Mass: 19097.180 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Homo sapiens (human) / References: GenBank: 1633547
#12: RNA chain modified H2a pre-mRNA


Mass: 16626.979 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Mus musculus (house mouse)

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Non-polymers , 1 types, 2 molecules

#13: Chemical ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Zn / Feature type: SUBJECT OF INVESTIGATION

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Details

Has ligand of interestY
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: The core of metazoan replication-dependent histone pre-mRNA 3'-end processing machinery
Type: COMPLEX / Entity ID: #1-#12 / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Trichoplusia ni (cabbage looper)
Buffer solutionpH: 7.5
Buffer componentConc.: 100 mM / Name: sodium chloride / Formula: NaCl
SpecimenConc.: 0.3 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: unspecified
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 295 K

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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 magnification: 22500 X / Nominal defocus max: 2500 nm / Nominal defocus min: 1200 nm / Calibrated defocus min: 900 nm / Calibrated defocus max: 2800 nm / Cs: 2.7 mm / C2 aperture diameter: 100 µm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recording
IDImaging-IDElectron dose (e/Å2)Detector modeFilm or detector model
1170COUNTINGGATAN K2 SUMMIT (4k x 4k)
2173GATAN K2 SUMMIT (4k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.17.1_3660: / Classification: refinement
EM software
IDNameCategory
4CTFFINDCTF correction
9PHENIXmodel refinement
13RELION3D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 325282 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00616902
ELECTRON MICROSCOPYf_angle_d1.00523070
ELECTRON MICROSCOPYf_dihedral_angle_d16.4976545
ELECTRON MICROSCOPYf_chiral_restr0.062681
ELECTRON MICROSCOPYf_plane_restr0.0072755

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