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- PDB-6qdv: Human post-catalytic P complex spliceosome -

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Entry
Database: PDB / ID: 6qdv
TitleHuman post-catalytic P complex spliceosome
Components
  • (Pre-mRNA-processing factor ...) x 2
  • (Pre-mRNA-splicing factor ...) x 6
  • (Small nuclear ribonucleoprotein ...SnRNP) x 6
  • (U2 small nuclear ribonucleoprotein ...) x 2
  • (U5 small nuclear ribonucleoprotein ...) x 2
  • 116 kDa U5 small nuclear ribonucleoprotein component
  • ATP-dependent RNA helicase DHX8
  • Cactin
  • Cell division cycle 5-like protein
  • Crooked neck-like protein 1
  • Eukaryotic initiation factor 4A-III
  • Intron lariat: MINX RNA
  • Intron-binding protein aquarius
  • Ligated exons: MINX mRNA
  • NF-kappa-B-activating protein
  • PRKR-interacting protein 1
  • Peptidyl-prolyl cis-trans isomerase-like 1
  • Pleiotropic regulator 1
  • Pre-mRNA-processing-splicing factor 8
  • Protein BUD31 homolog
  • Protein FAM32A
  • Protein mago nashi homolog 2
  • RNA-binding protein 8A
  • Replication stress response regulator SDE2
  • SNW domain-containing protein 1
  • Serine/arginine repetitive matrix protein 2
  • Small nuclear ribonucleoprotein-associated proteins B and B'
  • Spliceosome-associated protein CWC15 homolog
  • U2 snRNAU2 spliceosomal RNA
  • U5 snRNAU5 spliceosomal RNA
  • U6 snRNAU6 spliceosomal RNA
KeywordsSPLICING / spliceosome / RNA / complex
Function / homology
Function and homology information


ISG15 antiviral mechanism / SLBP independent Processing of Histone Pre-mRNAs / Transport of Mature mRNA derived from an Intron-Containing Transcript / mRNA Splicing - Major Pathway / Nonsense Mediated Decay (NMD) enhanced by the Exon Junction Complex (EJC) / NOTCH4 Intracellular Domain Regulates Transcription / NOTCH3 Intracellular Domain Regulates Transcription / Regulation of expression of SLITs and ROBOs / Gene and protein expression by JAK-STAT signaling after Interleukin-12 stimulation / RUNX3 regulates NOTCH signaling ...ISG15 antiviral mechanism / SLBP independent Processing of Histone Pre-mRNAs / Transport of Mature mRNA derived from an Intron-Containing Transcript / mRNA Splicing - Major Pathway / Nonsense Mediated Decay (NMD) enhanced by the Exon Junction Complex (EJC) / NOTCH4 Intracellular Domain Regulates Transcription / NOTCH3 Intracellular Domain Regulates Transcription / Regulation of expression of SLITs and ROBOs / Gene and protein expression by JAK-STAT signaling after Interleukin-12 stimulation / RUNX3 regulates NOTCH signaling / SLBP Dependent Processing of Replication-Dependent Histone Pre-mRNAs / RNA Polymerase II Transcription Termination / mRNA 3'-end processing / mRNA Splicing - Minor Pathway / Gap-filling DNA repair synthesis and ligation in TC-NER / Pre-NOTCH Transcription and Translation / Dual incision in TC-NER / Transcription-Coupled Nucleotide Excision Repair (TC-NER) / Formation of TC-NER Pre-Incision Complex / Deadenylation of mRNA / Notch-HLH transcription pathway / Constitutive Signaling by NOTCH1 HD+PEST Domain Mutants / Constitutive Signaling by NOTCH1 PEST Domain Mutants / SMAD2/SMAD3:SMAD4 heterotrimer regulates transcription / NOTCH1 Intracellular Domain Regulates Transcription / Regulation of gene expression in late stage (branching morphogenesis) pancreatic bud precursor cells / snRNP Assembly / negative regulation of selenocysteine insertion sequence binding / negative regulation of selenocysteine incorporation / positive regulation of vitamin D receptor signaling pathway / cellular response to selenite ion / granulocyte differentiation / cellular response to interleukin-2 / cellular response to wortmannin / negative regulation of interleukin-8 production / cellular response to prolactin / spliceosomal complex disassembly / regulation of cell cycle arrest / U2 snRNP binding / snRNP binding / U7 snRNA binding / exon-exon junction complex / histone pre-mRNA DCP binding / leucine zipper domain binding / U7 snRNP / pre-mRNA 3'-splice site binding / negative regulation of interferon-beta production / signal transduction involved in DNA damage checkpoint / histone pre-mRNA 3'end processing complex / negative regulation of excitatory postsynaptic potential / hematopoietic stem cell proliferation / exploration behavior / selenocysteine insertion sequence binding / methylosome / cis assembly of pre-catalytic spliceosome / mitotic G2 DNA damage checkpoint / go:0034459: / regulation of translation at postsynapse, modulating synaptic transmission / inner cell mass cell proliferation / histone mRNA metabolic process / regulation of mRNA splicing, via spliceosome / regulation of vitamin D receptor signaling pathway / U2-type catalytic step 1 spliceosome / telomerase holoenzyme complex / renal system process / positive regulation of neurogenesis / negative regulation of toll-like receptor signaling pathway / RNA splicing, via transesterification reactions / generation of catalytic spliceosome for first transesterification step / hair cycle / protein phosphatase 1 binding / spliceosome conformational change to release U4 (or U4atac) and U1 (or U11) / protein kinase inhibitor activity / negative regulation of phosphorylation / positive regulation of androgen receptor activity / transcription corepressor binding / RNA stem-loop binding / ubiquitin-ubiquitin ligase activity / mRNA 3'-splice site recognition / U2-type precatalytic spliceosome / U1 snRNP binding / WD40-repeat domain binding / alternative mRNA splicing, via spliceosome / positive regulation of mRNA splicing, via spliceosome / positive regulation of astrocyte differentiation / pICln-Sm protein complex / telomerase RNA binding / U4 snRNP / import into nucleus / small nuclear ribonucleoprotein complex / regulation of retinoic acid receptor signaling pathway / second spliceosomal transesterification activity / negative regulation of lipopolysaccharide-mediated signaling pathway / U5 snRNA binding / C2H2 zinc finger domain binding / post-mRNA release spliceosomal complex / retinoic acid receptor binding / SMN-Sm protein complex / Notch binding / U2 snRNA binding
Myb-like DNA-binding domain / Pre-mRNA-processing-splicing factor 8, U5-snRNA-binding / RNA-binding domain, S1 / PRP8 domain IV core / Pre-mRNA splicing factor component Cdc5p/Cef1 / Pre-mRNA-splicing factor SLU7 domain / Cyclophilin-type peptidyl-prolyl cis-trans isomerase, conserved site / Ribosomal protein S5 domain 2-type fold / G-protein beta WD-40 repeat / WD40 repeat, conserved site ...Myb-like DNA-binding domain / Pre-mRNA-processing-splicing factor 8, U5-snRNA-binding / RNA-binding domain, S1 / PRP8 domain IV core / Pre-mRNA splicing factor component Cdc5p/Cef1 / Pre-mRNA-splicing factor SLU7 domain / Cyclophilin-type peptidyl-prolyl cis-trans isomerase, conserved site / Ribosomal protein S5 domain 2-type fold / G-protein beta WD-40 repeat / WD40 repeat, conserved site / Tetratricopeptide repeat / RNA recognition motif, spliceosomal PrP8 / Pre-mRNA-processing-splicing factor 8, U6-snRNA-binding / Spt5 C-terminal domain / Cactin, C-terminal / Cactin, central domain / BUD31/G10-related, conserved site / WD40-repeat-containing domain / Myb domain / SKI-interacting protein, SKIP / Small ribonucleoprotein associated, SmB/SmN / Sm-like protein Lsm6/SmF / Armadillo-type fold / MIF4G-like domain superfamily / Cyclophilin-type peptidyl-prolyl cis-trans isomerase / Sde2, N-terminal ubiquitin domain / Immunoglobulin E-set / Small nuclear ribonucleoprotein Sm D3 / Winged helix DNA-binding domain superfamily / WD40-repeat-containing domain superfamily / RNA-binding domain superfamily / C2 domain superfamily / 116kDa U5 small nuclear ribonucleoprotein component, C-terminal / EF-G domain III/V-like / U2 small nuclear ribonucleoprotein B'', RNA recognition motif 1 / U2 small nuclear ribonucleoprotein B'', RNA recognition motif 2 / Small nuclear ribonucleoprotein D1 / Small nuclear ribonucleoprotein F / Small nuclear ribonucleoprotein G / CWF11 family / RBM8, RNA recognition motif / Pre-mRNA-processing factor 17 / Leucine-rich repeat domain superfamily / Intron-binding protein aquarius, N-terminal / 116kDa U5 small nuclear ribonucleoprotein component, N-terminal / Cyclophilin-like domain superfamily / Pre-mRNA-processing-splicing factor 8 / P-loop containing nucleoside triphosphate hydrolase / Small nuclear ribonucleoprotein Sm D2 / Like-Sm (LSM) domain containing protein, LSm4/SmD1/SmD3 / Small nuclear ribonucleoprotein E / WD40/YVTN repeat-like-containing domain superfamily / Ribosomal protein S5 domain 2-type fold, subgroup / Zinc finger, CCCH-type superfamily / DNA/RNA helicase, ATP-dependent, DEAH-box type, conserved site / Sec63 domain / Translation elongation factor EFTu-like, domain 2 / Mago nashi protein / SKI-interacting protein SKIP, SNW domain / Initiation factor eIF-4 gamma, MA3 / MIF4G-like, type 3 / U box domain / U2A'/phosphoprotein 32 family A, C-terminal / HAT (Half-A-TPR) repeat / S1 domain / Cyclophilin-type peptidyl-prolyl cis-trans isomerase domain / Translation elongation factor EFG/EF2, domain IV / G10 protein / WD40 repeat / Helicase, C-terminal / Leucine-rich repeat / LSM domain, eukaryotic/archaea-type / SANT/Myb domain / Transcription factor, GTP-binding domain / Ricin B, lectin domain / Elongation factor EFG, domain V-like / ATP-dependent RNA helicase DEAD-box, conserved site / Small GTP-binding protein domain / Pre-mRNA-splicing factor Cwf15/Cwc15 / RNA helicase, DEAD-box type, Q motif / PRO8NT domain / Helicase superfamily 1/2, ATP-binding domain / Pre-mRNA-splicing factor 19 / Protein FAM32A / mRNA splicing factor SYF2 / mRNA splicing factor Cwf21 domain / Zinc finger, RING/FYVE/PHD-type / Tetratricopeptide repeat-containing domain / PROCT domain / Nucleotide-binding alpha-beta plait domain superfamily / PROCN domain / Nucleic acid-binding, OB-fold / Helicase-associated domain / Ribonuclease H-like superfamily / Tetratricopeptide-like helical domain superfamily / Domain of unknown function DUF1605 / DEAD/DEAH box helicase domain / LSM domain superfamily / PRKR-interacting protein 1 / NF-kappa-B-activating protein, C-terminal
RNA-binding protein 8A / Crooked neck-like protein 1 / Pre-mRNA-processing-splicing factor 8 / gb:36515: / NF-kappa-B-activating protein / Cactin / Protein mago nashi homolog 2 / U5 small nuclear ribonucleoprotein 40 kDa protein / Cell division cycle 5-like protein / PRKR-interacting protein 1 ...RNA-binding protein 8A / Crooked neck-like protein 1 / Pre-mRNA-processing-splicing factor 8 / gb:36515: / NF-kappa-B-activating protein / Cactin / Protein mago nashi homolog 2 / U5 small nuclear ribonucleoprotein 40 kDa protein / Cell division cycle 5-like protein / PRKR-interacting protein 1 / Protein FAM32A / Replication stress response regulator SDE2 / Pre-mRNA-splicing factor SYF1 / Pre-mRNA-splicing factor RBM22 / Spliceosome-associated protein CWC15 homolog / Pre-mRNA-processing factor 19 / Serine/arginine repetitive matrix protein 2 / Peptidyl-prolyl cis-trans isomerase-like 1 / Pre-mRNA-splicing factor CWC22 homolog / ATP-dependent RNA helicase DHX8 / 116 kDa U5 small nuclear ribonucleoprotein component / U2 small nuclear ribonucleoprotein B'' / Pleiotropic regulator 1 / RNA helicase aquarius / Pre-mRNA-processing factor 17 / U5 small nuclear ribonucleoprotein 200 kDa helicase / Pre-mRNA-splicing factor SPF27 / Pre-mRNA-splicing factor SLU7 / Pre-mRNA-splicing factor SYF2 / U2 small nuclear ribonucleoprotein A' / SNW domain-containing protein 1 / Small nuclear ribonucleoprotein-associated proteins B and B' / Eukaryotic initiation factor 4A-III / Protein BUD31 homolog / 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
Biological speciesHomo sapiens (human)
Human adenovirus 2
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsFica, S.M. / Oubridge, C. / Wilkinson, M.E. / Newman, A.J. / Nagai, K.
Funding supportUnited Kingdom , 2件
OrganizationGrant numberCountry
Medical Research Council (United Kingdom)MC_U105184330United Kingdom
European Research CouncilAdG-693087-SPLICE3D
CitationJournal: Science / Year: 2019
Title: A human postcatalytic spliceosome structure reveals essential roles of metazoan factors for exon ligation.
Authors: Sebastian M Fica / Chris Oubridge / Max E Wilkinson / Andrew J Newman / Kiyoshi Nagai /
Abstract: During exon ligation, the spliceosome recognizes the 3'-splice site (3'SS) of precursor messenger RNA (pre-mRNA) through non-Watson-Crick pairing with the 5'SS and the branch adenosine, in a ...During exon ligation, the spliceosome recognizes the 3'-splice site (3'SS) of precursor messenger RNA (pre-mRNA) through non-Watson-Crick pairing with the 5'SS and the branch adenosine, in a conformation stabilized by Prp18 and Prp8. Here we present the 3.3-angstrom cryo-electron microscopy structure of a human postcatalytic spliceosome just after exon ligation. The 3'SS docks at the active site through conserved RNA interactions in the absence of Prp18. Unexpectedly, the metazoan-specific FAM32A directly bridges the 5'-exon and intron 3'SS of pre-mRNA and promotes exon ligation, as shown by functional assays. CACTIN, SDE2, and NKAP-factors implicated in alternative splicing-further stabilize the catalytic conformation of the spliceosome during exon ligation. Together these four proteins act as exon ligation factors. Our study reveals how the human spliceosome has co-opted additional proteins to modulate a conserved RNA-based mechanism for 3'SS selection and to potentially fine-tune alternative splicing at the exon ligation stage.
Validation Report
SummaryFull reportAbout validation report
DateDeposition: Jan 3, 2019 / Release: Feb 20, 2019
RevisionDateData content typeGroupCategoryItemProviderType
1.0Feb 20, 2019Structure modelrepositoryInitial release
1.1Feb 27, 2019Structure modelData collection / Database referencescitation / citation_author / database_PDB_rev / database_PDB_rev_record / em_admin / pdbx_database_proc / pdbx_seq_map_depositor_info_citation.journal_volume / _citation.page_first / _citation.page_last / _citation_author.identifier_ORCID / _em_admin.last_update / _pdbx_seq_map_depositor_info.one_letter_code / _pdbx_seq_map_depositor_info.one_letter_code_mod

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Assembly

Deposited unit
2: U2 snRNA
5: U5 snRNA
6: U6 snRNA
7: Eukaryotic initiation factor 4A-III
8: RNA-binding protein 8A
9: Protein mago nashi homolog 2
A: Pre-mRNA-processing-splicing factor 8
B: U5 small nuclear ribonucleoprotein 200 kDa helicase
C: 116 kDa U5 small nuclear ribonucleoprotein component
D: PRKR-interacting protein 1
E: Ligated exons: MINX mRNA
F: Cactin
G: Protein FAM32A
H: Pre-mRNA-splicing factor CWC22 homolog
I: Intron lariat: MINX RNA
J: Pleiotropic regulator 1
K: SNW domain-containing protein 1
L: Protein BUD31 homolog
M: Pre-mRNA-splicing factor RBM22
N: U5 small nuclear ribonucleoprotein 40 kDa protein
O: Cell division cycle 5-like protein
P: Spliceosome-associated protein CWC15 homolog
R: Serine/arginine repetitive matrix protein 2
S: Crooked neck-like protein 1
T: Pre-mRNA-splicing factor SYF1
U: Intron-binding protein aquarius
V: ATP-dependent RNA helicase DHX8
W: U2 small nuclear ribonucleoprotein A'
Y: U2 small nuclear ribonucleoprotein B''
Z: NF-kappa-B-activating protein
b: Small nuclear ribonucleoprotein-associated proteins B and B'
c: Pre-mRNA-splicing factor SLU7
d: Small nuclear ribonucleoprotein Sm D3
e: Small nuclear ribonucleoprotein E
f: Small nuclear ribonucleoprotein F
g: Small nuclear ribonucleoprotein G
h: Small nuclear ribonucleoprotein Sm D1
i: Peptidyl-prolyl cis-trans isomerase-like 1
j: Small nuclear ribonucleoprotein Sm D2
k: Small nuclear ribonucleoprotein-associated proteins B and B'
l: Small nuclear ribonucleoprotein Sm D1
m: Small nuclear ribonucleoprotein Sm D2
n: Small nuclear ribonucleoprotein Sm D3
o: Pre-mRNA-processing factor 17
p: Small nuclear ribonucleoprotein E
q: Small nuclear ribonucleoprotein F
r: Small nuclear ribonucleoprotein G
s: Pre-mRNA-splicing factor SPF27
t: Pre-mRNA-processing factor 19
u: Pre-mRNA-processing factor 19
v: Pre-mRNA-processing factor 19
w: Pre-mRNA-processing factor 19
y: Pre-mRNA-splicing factor SYF2
z: Replication stress response regulator SDE2
hetero molecules


Theoretical massNumber of molelcules
Total (without water)2,322,14372
Polymers2,319,78654
Non-polymers2,35718
Water0
1


  • Idetical with deposited unit
  • defined by author
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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RNA chain , 5 types, 5 molecules 256EI

#1: RNA chain U2 snRNA / U2 spliceosomal RNA


Mass: 60459.621 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human)
#2: RNA chain U5 snRNA / U5 spliceosomal RNA


Mass: 36908.668 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: GenBank: 36515
#3: RNA chain U6 snRNA / U6 spliceosomal RNA


Mass: 34098.270 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human)
#11: RNA chain Ligated exons: MINX mRNA


Mass: 4437.700 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Human adenovirus 2
#15: RNA chain Intron lariat: MINX RNA


Mass: 27772.039 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Human adenovirus 2

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Protein/peptide , 21 types, 22 molecules 789ACDFGJKLOPRSUVZbkiz

#4: Protein/peptide Eukaryotic initiation factor 4A-III / eIF4A-III / ATP-dependent RNA helicase DDX48 / ATP-dependent RNA helicase eIF4A-3 / DEAD box protein 48 / Eukaryotic initiation factor 4A-like NUK-34 / Eukaryotic translation initiation factor 4A isoform 3 / Nuclear matrix protein 265 / hNMP 265


Mass: 44691.359 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: P38919, EC: 3.6.4.13
#5: Protein/peptide RNA-binding protein 8A / Binder of OVCA1-1 / BOV-1 / RNA-binding motif protein 8A / RNA-binding protein Y14 / Ribonucleoprotein RBM8A


Mass: 10370.526 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q9Y5S9
#6: Protein/peptide Protein mago nashi homolog 2


Mass: 16929.314 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q96A72
#7: Protein/peptide Pre-mRNA-processing-splicing factor 8 / 220 kDa U5 snRNP-specific protein / PRP8 homolog / Splicing factor Prp8 / p220


Mass: 273974.250 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q6P2Q9
#9: Protein/peptide 116 kDa U5 small nuclear ribonucleoprotein component / Elongation factor Tu GTP-binding domain-containing protein 2 / SNU114 homolog / hSNU114 / U5 snRNP-specific protein / 116 kDa / U5-116 kDa


Mass: 101231.938 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q15029
#10: Protein/peptide PRKR-interacting protein 1


Mass: 14476.882 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q9H875
#12: Protein/peptide Cactin / / Renal carcinoma antigen NY-REN-24


Mass: 15210.249 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q8WUQ7
#13: Protein/peptide Protein FAM32A / Ovarian tumor-associated gene 12 / OTAG-12


Mass: 7269.374 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q9Y421
#16: Protein/peptide Pleiotropic regulator 1


Mass: 35782.395 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: O43660
#17: Protein/peptide SNW domain-containing protein 1 / Nuclear protein SkiP / Nuclear receptor coactivator NCoA-62 / Ski-interacting protein


Mass: 33819.922 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q13573
#18: Protein/peptide Protein BUD31 homolog / Protein EDG-2 / Protein G10 homolog


Mass: 17032.850 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: P41223
#21: Protein/peptide Cell division cycle 5-like protein / Cdc5-like protein / Pombe cdc5-related protein


Mass: 92406.883 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q99459
#22: Protein/peptide Spliceosome-associated protein CWC15 homolog


Mass: 26674.447 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q9P013
#23: Protein/peptide Serine/arginine repetitive matrix protein 2 / 300 kDa nuclear matrix antigen / Serine/arginine-rich splicing factor-related nuclear matrix protein of 300 kDa / Ser/Arg-related nuclear matrix protein of 300 kDa / Splicing coactivator subunit SRm300 / Tax-responsive enhancer element-binding protein 803 / TaxREB803


Mass: 2767.128 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q9UQ35
#24: Protein/peptide Crooked neck-like protein 1 / Crooked neck homolog / hCrn


Mass: 100610.008 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q9BZJ0
#26: Protein/peptide Intron-binding protein aquarius / Intron-binding protein of 160 kDa / IBP160


Mass: 171502.453 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: O60306, EC: 3.6.4.13
#27: Protein/peptide ATP-dependent RNA helicase DHX8 / DEAH box protein 8 / RNA helicase HRH1


Mass: 139522.312 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: DHX8, DDX8 / Production host: Saccharomyces cerevisiae (baker's yeast) / References: UniProt: Q14562, EC: 3.6.4.13
#30: Protein/peptide NF-kappa-B-activating protein /


Mass: 3321.766 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q8N5F7
#31: Protein/peptide Small nuclear ribonucleoprotein-associated proteins B and B' / snRNP-B / Sm protein B/B' / SmB/B'


Mass: 9551.284 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: P14678
#38: Protein/peptide Peptidyl-prolyl cis-trans isomerase-like 1 / PPIase / Rotamase PPIL1


Mass: 18055.529 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q9Y3C6, EC: 5.2.1.8
#44: Protein/peptide Replication stress response regulator SDE2


Mass: 4036.538 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q6IQ49

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U5 small nuclear ribonucleoprotein ... , 2 types, 2 molecules BN

#8: Protein/peptide U5 small nuclear ribonucleoprotein 200 kDa helicase / Activating signal cointegrator 1 complex subunit 3-like 1 / BRR2 homolog / U5 snRNP-specific 200 kDa protein / U5-200KD


Mass: 197053.984 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: O75643, EC: 3.6.4.13
#20: Protein/peptide U5 small nuclear ribonucleoprotein 40 kDa protein / U5-40K / 38 kDa-splicing factor / Prp8-binding protein / hPRP8BP / U5 snRNP-specific 40 kDa protein / WD repeat-containing protein 57


Mass: 34092.410 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q96DI7

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Pre-mRNA-splicing factor ... , 6 types, 6 molecules HMTcsy

#14: Protein/peptide Pre-mRNA-splicing factor CWC22 homolog / Nucampholin homolog / fSAPb


Mass: 105646.578 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q9HCG8
#19: Protein/peptide Pre-mRNA-splicing factor RBM22 / RNA-binding motif protein 22 / Zinc finger CCCH domain-containing protein 16


Mass: 33156.129 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q9NW64
#25: Protein/peptide Pre-mRNA-splicing factor SYF1 / Protein HCNP / XPA-binding protein 2


Mass: 100148.711 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q9HCS7
#32: Protein/peptide Pre-mRNA-splicing factor SLU7 / hSlu7


Mass: 68510.844 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: O95391
#41: Protein/peptide Pre-mRNA-splicing factor SPF27 / Breast carcinoma-amplified sequence 2 / DNA amplified in mammary carcinoma 1 protein / Spliceosome-associated protein SPF 27


Mass: 26163.420 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: O75934
#43: Protein/peptide Pre-mRNA-splicing factor SYF2 / CCNDBP1-interactor / p29


Mass: 17302.371 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: O95926

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

#28: Protein/peptide U2 small nuclear ribonucleoprotein A' / U2 snRNP A'


Mass: 18697.730 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: P09661
#29: Protein/peptide U2 small nuclear ribonucleoprotein B'' / U2 snRNP B''


Mass: 10688.563 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, 12 molecules dnepfqgrhljm

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


Mass: 9460.979 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: P62318
#34: Protein/peptide Small nuclear ribonucleoprotein E / snRNP-E / Sm protein E / SmE


Mass: 9582.185 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: P62304
#35: Protein/peptide Small nuclear ribonucleoprotein F / snRNP-F / Sm protein F / SmF


Mass: 8038.415 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: P62306
#36: Protein/peptide Small nuclear ribonucleoprotein G / snRNP-G / Sm protein G / SmG


Mass: 8160.629 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: P62308
#37: Protein/peptide Small nuclear ribonucleoprotein Sm D1 / Sm-D1 / Sm-D autoantigen / snRNP core protein D1


Mass: 9086.731 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: P62314
#39: Protein/peptide Small nuclear ribonucleoprotein Sm D2 / Sm-D2 / snRNP core protein D2


Mass: 13551.928 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: P62316

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Pre-mRNA-processing factor ... , 2 types, 5 molecules otuvw

#40: Protein/peptide Pre-mRNA-processing factor 17 / Cell division cycle 40 homolog / EH-binding protein 3 / Ehb3 / PRP17 homolog / hPRP17


Mass: 59116.055 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: O60508
#42: Protein/peptide
Pre-mRNA-processing factor 19 / Nuclear matrix protein 200 / PRP19/PSO4 homolog / hPso4 / RING-type E3 ubiquitin transferase PRP19 / Senescence evasion factor


Mass: 55245.547 Da / Num. of mol.: 4 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q9UMS4, EC: 2.3.2.27

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Non-polymers , 6 types, 18 molecules

#45: Chemical
ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 7 / Source method: obtained synthetically / Formula: Mg / Magnesium
#46: Chemical ChemComp-K / POTASSIUM ION


Mass: 39.098 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: K / Potassium
#47: Chemical ChemComp-ATP / ADENOSINE-5'-TRIPHOSPHATE


Mass: 507.181 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C10H16N5O13P3 / Adenosine triphosphate / Comment: ATP (energy-carrying molecule) *YM
#48: Chemical ChemComp-GTP / GUANOSINE-5'-TRIPHOSPHATE


Mass: 523.180 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C10H16N5O14P3 / Guanosine triphosphate / Comment: GTP (energy-carrying molecule) *YM
#49: Chemical
ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 7 / Source method: obtained synthetically / Formula: Zn / Zinc
#50: Chemical ChemComp-I6P / INOSITOL 1,2,3,4,5,6-HEXAKISPHOSPHATE / MYO-INOSITOL HEXAKISPHOSPHATE


Mass: 660.035 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C6H18O24P6 / Phytic acid

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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

Component

Type: COMPLEX

IDNameEntity IDParent-IDSource
1Human post-catalytic P complex spliceosome1,2,3,4,5,6,7,8,9,10,11,12,13,14,15,16,17,18,19,20,21,22,23,24,25,26,27,28,29,30,31,32,33,34,35,36,37,38,39,40,41,42,43,440MULTIPLE SOURCES
2Human post-catalytic P complex spliceosome1,2,3,4,5,6,7,8,9,10,12,13,14,16,17,18,19,20,21,22,23,24,25,26,28,29,30,31,32,33,34,35,36,37,38,39,40,41,42,43,441NATURAL
3ATP-dependent RNA helicase DHX8271RECOMBINANT
4MINX introns and exons11,151RECOMBINANT
Molecular weightValue: 3.0 MDa / Experimental value: NO
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
12Homo sapiens (human)9606
23Homo sapiens (human)9606
34Human adenovirus 210515
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-ID
13Saccharomyces cerevisiae (baker's yeast)4932
24synthetic construct (others)32630
Buffer solutionpH: 7.9
Buffer component

Buffer-ID: 1

IDConc.NameFormula
120 mMHEPES
2100 mMKCl
30.2 mMEDTAEthylenediaminetetraacetic acid
SpecimenConc.: 0.3 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid type: Quantifoil R2/2
VitrificationInstrument: FEI VITROBOT MARK III / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K
Details: 3 uL sample was applied to the grid, left for 25s, then blotted for 2.5-3.5s and immediately plunged into liquid 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 magnification: 135000 X / Nominal defocus max: 3500 nm / Nominal defocus min: 1500 nm / Cs: 2.7 mm
Image recordingAverage exposure time: 10 sec. / Electron dose: 53 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 2 / Num. of real images: 6200
Image scansMovie frames/image: 40 / Used frames/image: 1-40

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Processing

EM software
IDNameVersionCategory
2EPUimage acquisition
9PHENIX1.14model refinement
10RELION3initial Euler assignment
11RELION3final Euler assignment
13RELION33D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 103860 / Symmetry type: POINT

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