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- PDB-8hk1: The cryo-EM structure of human pre-17S U2 snRNP -

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Entry
Database: PDB / ID: 8hk1
TitleThe cryo-EM structure of human pre-17S U2 snRNP
Components
  • (Small nuclear ribonucleoprotein ...SnRNP) x 6
  • (Splicing factor 3A subunit ...) x 3
  • (Splicing factor 3B subunit ...) x 5
  • (U2 small nuclear ribonucleoprotein ...) x 2
  • ATP-dependent RNA helicase DDX42
  • HIV Tat-specific factor 1
  • PHD finger-like domain-containing protein 5A
  • Small nuclear ribonucleoprotein-associated proteins B and B'
  • U2 snRNAU2 spliceosomal RNA
KeywordsSPLICING / DDX42 / U2 snRNP / spliceosome
Function / homology
Function and homology information


U11/U12 snRNP / chromatin-protein adaptor activity / snRNP binding / U2 snRNP binding / U7 snRNA binding / histone pre-mRNA DCP binding / U7 snRNP / B-WICH complex / histone pre-mRNA 3'end processing complex / SLBP independent Processing of Histone Pre-mRNAs ...U11/U12 snRNP / chromatin-protein adaptor activity / snRNP binding / U2 snRNP binding / U7 snRNA binding / histone pre-mRNA DCP binding / U7 snRNP / B-WICH complex / histone pre-mRNA 3'end processing complex / SLBP independent Processing of Histone Pre-mRNAs / SLBP Dependent Processing of Replication-Dependent Histone Pre-mRNAs / protein localization to site of double-strand break / splicing factor binding / protein methylation / U12-type spliceosomal complex / methylosome / 7-methylguanosine cap hypermethylation / U1 snRNP binding / pICln-Sm protein complex / poly-ADP-D-ribose modification-dependent protein binding / RNA splicing, via transesterification reactions / small nuclear ribonucleoprotein complex / P granule / SMN-Sm protein complex / spliceosomal tri-snRNP complex / telomerase RNA binding / positive regulation of mRNA splicing, via spliceosome / : / telomerase holoenzyme complex / mRNA cis splicing, via spliceosome / U2-type spliceosomal complex / U2-type precatalytic spliceosome / commitment complex / U2-type prespliceosome assembly / U2-type catalytic step 2 spliceosome / U4 snRNP / SAGA complex / U2 snRNP / RNA Polymerase II Transcription Termination / positive regulation of transcription by RNA polymerase III / U1 snRNP / U2-type prespliceosome / precatalytic spliceosome / spliceosomal complex assembly / positive regulation of transcription by RNA polymerase I / mRNA Splicing - Minor Pathway / regulation of RNA splicing / mRNA 3'-splice site recognition / U5 snRNP / U2 snRNA binding / Cajal body / spliceosomal snRNP assembly / regulation of DNA repair / U1 snRNA binding / U4/U6 x U5 tri-snRNP complex / Gene and protein expression by JAK-STAT signaling after Interleukin-12 stimulation / catalytic step 2 spliceosome / mRNA Splicing - Major Pathway / RNA splicing / stem cell differentiation / double-strand break repair via homologous recombination / spliceosomal complex / protein localization / B-WICH complex positively regulates rRNA expression / negative regulation of protein catabolic process / cytoplasmic ribonucleoprotein granule / fibrillar center / mRNA processing / mRNA splicing, via spliceosome / positive regulation of neuron projection development / nuclear matrix / site of double-strand break / snRNP Assembly / SARS-CoV-2 modulates host translation machinery / spermatogenesis / regulation of apoptotic process / RNA helicase activity / nuclear body / RNA helicase / nuclear speck / chromatin remodeling / mRNA binding / protein-containing complex binding / nucleolus / positive regulation of DNA-templated transcription / enzyme binding / ATP hydrolysis activity / positive regulation of transcription by RNA polymerase II / DNA binding / RNA binding / extracellular exosome / zinc ion binding / nucleoplasm / ATP binding / membrane / nucleus / cytosol / cytoplasm
Similarity search - Function
TatSF1-like, RNA recognition motif 1 / TatSF1-like / SF3B4, RNA recognition motif 2 / Splicing factor SF3a60 binding domain / Splicing factor SF3a60 binding domain / : / Replication stress response SDE2 C-terminal / Cactus-binding C-terminus of cactin protein / Splicing factor 3A subunit 1, ubiquitin domain / U2 small nuclear ribonucleoprotein B'', RNA recognition motif 2 ...TatSF1-like, RNA recognition motif 1 / TatSF1-like / SF3B4, RNA recognition motif 2 / Splicing factor SF3a60 binding domain / Splicing factor SF3a60 binding domain / : / Replication stress response SDE2 C-terminal / Cactus-binding C-terminus of cactin protein / Splicing factor 3A subunit 1, ubiquitin domain / U2 small nuclear ribonucleoprotein B'', RNA recognition motif 2 / U2 small nuclear ribonucleoprotein B'', RNA recognition motif 1 / SF3A2 domain / Pre-mRNA-splicing factor SF3a complex subunit 2 (Prp11) / Splicing factor SF3a60 /Prp9 subunit, C-terminal / SF3A3 domain / SF3B4, RNA recognition motif 1 / SF3a60/Prp9 C-terminal / Pre-mRNA-splicing factor SF3A3, of SF3a complex, Prp9 / Splicing factor 3B, subunit 5 / Splicing factor 3A subunit 1 / Splicing factor 3B subunit 1 / Splicing factor 3B subunit 1 / Splicing factor 3A subunit 1, conserved domain / Pre-mRNA splicing factor PRP21 like protein / Domain of unknown function DUF382 / Domain of unknown function (DUF382) / SWAP/Surp / SWAP/Surp superfamily / Surp module / Zinc-finger of C2H2 type / SURP motif repeat profile. / Suppressor-of-White-APricot splicing regulator / PHF5-like / PHF5-like protein / PSP, proline-rich / PSP / proline-rich domain in spliceosome associated proteins / Splicing factor 3B subunit 5/RDS3 complex subunit 10 / Splicing factor 3B subunit 10 (SF3b10) / Small ribonucleoprotein associated, SmB/SmN / Splicing factor 3B subunit 1-like / Matrin/U1-C, C2H2-type zinc finger / Zinc finger matrin-type profile. / Small nuclear ribonucleoprotein D1 / U2A'/phosphoprotein 32 family A, C-terminal / occurring C-terminal to leucine-rich repeats / Leucine-rich repeat / SAP domain / SAP motif profile. / Putative DNA-binding (bihelical) motif predicted to be involved in chromosomal organisation / SAP domain / Small nuclear ribonucleoprotein Sm D3 / Small nuclear ribonucleoprotein Sm D2 / 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 / Matrin/U1-C-like, C2H2-type zinc finger / U1-like zinc finger / Sm-like protein Lsm6/SmF / LSM domain / LSM domain, eukaryotic/archaea-type / snRNP Sm proteins / Cleavage/polyadenylation specificity factor, A subunit, N-terminal / Mono-functional DNA-alkylating methyl methanesulfonate N-term / Cleavage/polyadenylation specificity factor, A subunit, C-terminal / CPSF A subunit region / : / Sm domain profile. / DEAD-box subfamily ATP-dependent helicases signature. / ATP-dependent RNA helicase DEAD-box, conserved site / RNA helicase, DEAD-box type, Q motif / DEAD-box RNA helicase Q motif profile. / LSM domain superfamily / Zinc finger C2H2 superfamily / Leucine-rich repeat profile. / DEAD/DEAH box helicase / DEAD/DEAH box helicase domain / Zinc finger C2H2-type / Leucine-rich repeat / RNA recognition motif / RNA recognition motif / Eukaryotic RNA Recognition Motif (RRM) profile. / RNA recognition motif domain / Helicase conserved C-terminal domain / Leucine-rich repeat domain superfamily / RNA-binding domain superfamily / Armadillo-like helical / Ubiquitin family / Ubiquitin homologues / Ubiquitin-like domain / Ubiquitin domain profile. / helicase superfamily c-terminal domain / Superfamilies 1 and 2 helicase C-terminal domain profile. / Superfamilies 1 and 2 helicase ATP-binding type-1 domain profile. / DEAD-like helicases superfamily / Helicase, C-terminal / Helicase superfamily 1/2, ATP-binding domain / Armadillo-type fold
Similarity search - Domain/homology
Chem-9B0 / : / RNA / RNA (> 10) / RNA (> 100) / 17S U2 SnRNP complex component HTATSF1 / Splicing factor 3B subunit 1 / U2 small nuclear ribonucleoprotein B'' / U2 small nuclear ribonucleoprotein A' / Small nuclear ribonucleoprotein-associated proteins B and B' ...Chem-9B0 / : / RNA / RNA (> 10) / RNA (> 100) / 17S U2 SnRNP complex component HTATSF1 / Splicing factor 3B subunit 1 / U2 small nuclear ribonucleoprotein B'' / U2 small nuclear ribonucleoprotein A' / Small nuclear ribonucleoprotein-associated proteins B and B' / Small nuclear ribonucleoprotein E / Small nuclear ribonucleoprotein F / Small nuclear ribonucleoprotein G / Small nuclear ribonucleoprotein Sm D1 / Small nuclear ribonucleoprotein Sm D2 / Small nuclear ribonucleoprotein Sm D3 / Splicing factor 3A subunit 3 / Splicing factor 3B subunit 2 / Splicing factor 3B subunit 3 / Splicing factor 3B subunit 4 / Splicing factor 3A subunit 2 / Splicing factor 3A subunit 1 / PHD finger-like domain-containing protein 5A / ATP-dependent RNA helicase DDX42 / Splicing factor 3B subunit 5
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.7 Å
AuthorsZhang, X. / Zhan, X. / Shi, Y.
Funding support China, 1items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC) China
CitationJournal: Nat Commun / Year: 2023
Title: Mechanisms of the RNA helicases DDX42 and DDX46 in human U2 snRNP assembly.
Authors: Fenghua Yang / Tong Bian / Xiechao Zhan / Zhe Chen / Zhihan Xing / Nicolas A Larsen / Xiaofeng Zhang / Yigong Shi /
Abstract: Three RNA helicases - DDX42, DDX46 and DHX15 - are found to be associated with human U2 snRNP, but their roles and mechanisms in U2 snRNP and spliceosome assembly are insufficiently understood. Here ...Three RNA helicases - DDX42, DDX46 and DHX15 - are found to be associated with human U2 snRNP, but their roles and mechanisms in U2 snRNP and spliceosome assembly are insufficiently understood. Here we report the cryo-electron microscopy (cryo-EM) structures of the DDX42-SF3b complex and a putative assembly precursor of 17S U2 snRNP that contains DDX42 (DDX42-U2 complex). DDX42 is anchored on SF3B1 through N-terminal sequences, with its N-plug occupying the RNA path of SF3B1. The binding mode of DDX42 to SF3B1 is in striking analogy to that of DDX46. In the DDX42-U2 complex, the N-terminus of DDX42 remains anchored on SF3B1, but the helicase domain has been displaced by U2 snRNA and TAT-SF1. Through in vitro assays, we show DDX42 and DDX46 are mutually exclusive in terms of binding to SF3b. Cancer-driving mutations of SF3B1 target the residues in the RNA path that directly interact with DDX42 and DDX46. These findings reveal the distinct roles of DDX42 and DDX46 in assembly of 17S U2 snRNP and provide insights into the mechanisms of SF3B1 cancer mutations.
History
DepositionNov 24, 2022Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Mar 8, 2023Provider: repository / Type: Initial release
Revision 1.1Aug 16, 2023Group: Data collection / Category: chem_comp_atom / chem_comp_bond

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
H: U2 snRNA
1: Splicing factor 3B subunit 1
2: Splicing factor 3B subunit 2
3: Splicing factor 3B subunit 3
4: Splicing factor 3B subunit 4
5: Splicing factor 3B subunit 5
6: PHD finger-like domain-containing protein 5A
A: Splicing factor 3A subunit 1
B: Splicing factor 3A subunit 2
C: Splicing factor 3A subunit 3
D: HIV Tat-specific factor 1
E: ATP-dependent RNA helicase DDX42
F: U2 small nuclear ribonucleoprotein A'
G: U2 small nuclear ribonucleoprotein B''
a: Small nuclear ribonucleoprotein Sm D2
b: Small nuclear ribonucleoprotein F
c: Small nuclear ribonucleoprotein E
d: Small nuclear ribonucleoprotein G
e: Small nuclear ribonucleoprotein Sm D3
f: Small nuclear ribonucleoprotein-associated proteins B and B'
g: Small nuclear ribonucleoprotein Sm D1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)1,045,13726
Polymers1,044,15621
Non-polymers9815
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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RNA chain , 1 types, 1 molecules H

#1: RNA chain U2 snRNA / U2 spliceosomal RNA


Mass: 60186.445 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: GenBank: 340097

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Splicing factor 3B subunit ... , 5 types, 5 molecules 12345

#2: Protein Splicing factor 3B subunit 1 / Pre-mRNA-splicing factor SF3b 155 kDa subunit / SF3b155 / Spliceosome-associated protein 155 / SAP 155


Mass: 146024.938 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: O75533
#3: Protein Splicing factor 3B subunit 2 / Pre-mRNA-splicing factor SF3b 145 kDa subunit / SF3b145 / Spliceosome-associated protein 145 / SAP 145


Mass: 100377.812 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q13435
#4: Protein Splicing factor 3B subunit 3 / Pre-mRNA-splicing factor SF3b 130 kDa subunit / SF3b130 / STAF130 / Spliceosome-associated protein ...Pre-mRNA-splicing factor SF3b 130 kDa subunit / SF3b130 / STAF130 / Spliceosome-associated protein 130 / SAP 130


Mass: 135718.844 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q15393
#5: Protein Splicing factor 3B subunit 4 / Pre-mRNA-splicing factor SF3b 49 kDa subunit / Spliceosome-associated protein 49 / SAP 49


Mass: 44436.570 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q15427
#6: Protein Splicing factor 3B subunit 5 / SF3b5 / Pre-mRNA-splicing factor SF3b 10 kDa subunit


Mass: 10149.369 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q9BWJ5

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Protein , 4 types, 4 molecules 6DEf

#7: Protein PHD finger-like domain-containing protein 5A / PHD finger-like domain protein 5A / Splicing factor 3B-associated 14 kDa protein / SF3b14b


Mass: 12427.524 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q7RTV0
#11: Protein HIV Tat-specific factor 1 / Tat-SF1


Mass: 85965.961 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: O43719
#12: Protein ATP-dependent RNA helicase DDX42 / DEAD box protein 42 / RNA helicase-like protein / RHELP / RNA helicase-related protein / RNAHP / ...DEAD box protein 42 / RNA helicase-like protein / RHELP / RNA helicase-related protein / RNAHP / SF3b DEAD box protein / Splicing factor 3B-associated 125 kDa protein / SF3b125


Mass: 103129.477 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q86XP3, RNA helicase
#20: Protein Small nuclear ribonucleoprotein-associated proteins B and B' / snRNP-B / Sm protein B/B' / SmB/B'


Mass: 24642.131 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: P14678

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Splicing factor 3A subunit ... , 3 types, 3 molecules ABC

#8: Protein Splicing factor 3A subunit 1 / SF3a120 / Spliceosome-associated protein 114 / SAP 114


Mass: 88991.094 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q15459
#9: Protein Splicing factor 3A subunit 2 / SF3a66 / Spliceosome-associated protein 62 / SAP 62


Mass: 49327.355 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q15428
#10: Protein Splicing factor 3A subunit 3 / SF3a60 / Spliceosome-associated protein 61 / SAP 61


Mass: 58934.844 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q12874

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U2 small nuclear ribonucleoprotein ... , 2 types, 2 molecules FG

#13: Protein U2 small nuclear ribonucleoprotein A' / U2 snRNP A'


Mass: 28456.584 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: P09661
#14: Protein U2 small nuclear ribonucleoprotein B'' / U2 snRNP B''


Mass: 25524.367 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: P08579

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Small nuclear ribonucleoprotein ... , 6 types, 6 molecules abcdeg

#15: Protein Small nuclear ribonucleoprotein Sm D2 / Sm-D2 / snRNP core protein D2


Mass: 13551.928 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: P62316
#16: Protein Small nuclear ribonucleoprotein F / snRNP-F / Sm protein F / SmF


Mass: 9734.171 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: P62306
#17: Protein Small nuclear ribonucleoprotein E / snRNP-E / Sm protein E / SmE


Mass: 10817.601 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: P62304
#18: Protein Small nuclear ribonucleoprotein G / snRNP-G / Sm protein G / SmG


Mass: 8508.084 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: P62308
#19: Protein Small nuclear ribonucleoprotein Sm D3 / Sm-D3 / snRNP core protein D3


Mass: 13940.308 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: P62318
#21: Protein Small nuclear ribonucleoprotein Sm D1 / Sm-D1 / Sm-D autoantigen / snRNP core protein D1


Mass: 13310.653 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: P62314

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Non-polymers , 2 types, 5 molecules

#22: Chemical ChemComp-9B0 / [(2~{S},3~{S},4~{E},6~{S},7~{R},10~{R})-3,7-dimethyl-2-[(2~{E},4~{E},6~{R})-6-methyl-6-oxidanyl-7-[(2~{R},3~{R})-3-[(2~{R},3~{S})-3-oxidanylpentan-2-yl]oxiran-2-yl]hepta-2,4-dien-2-yl]-7,10-bis(oxidanyl)-12-oxidanylidene-1-oxacyclododec-4-en-6-yl] 4-cycloheptylpiperazine-1-carboxylate


Mass: 718.960 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C40H66N2O9
#23: Chemical
ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: Zn

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Details

Has ligand of interestN

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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 human pre-17S U2 snRNP / Type: COMPLEX / Entity ID: #1-#21 / Source: NATURAL
Source (natural)Organism: Homo sapiens (human)
Buffer solutionpH: 7.9
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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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 FIELDBright-field microscopy / Nominal defocus max: 2000 nm / Nominal defocus min: 1500 nm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

CTF correctionType: NONE
3D reconstructionResolution: 2.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 430443 / Symmetry type: POINT

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