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- PDB-9dtr: Structure of the yeast post-catalytic P complex spliceosome at 2.... -

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Basic information

Entry
Database: PDB / ID: 9dtr
TitleStructure of the yeast post-catalytic P complex spliceosome at 2.3 Angstrom resolution
Components
  • (Pre-mRNA-processing factor ...) x 2
  • (Pre-mRNA-splicing factor ...) x 19
  • (Small nuclear ribonucleoprotein ...) x 6
  • (U2 small nuclear ribonucleoprotein ...) x 2
  • Pre-mRNA-processing protein 45
  • Protein FYV6
  • Small nuclear ribonucleoprotein-associated protein B
  • U2 snRNA
  • U5 snRNA
  • U6 snRNA
  • UBC4 lariat-intron
  • UBC4 mRNA spliced exons
KeywordsSPLICING / spliceosome / snRNA / yeast / ribonucleoprotein complex
Function / homology
Function and homology information


U2-type post-spliceosomal complex / spliceosomal complex disassembly / post-spliceosomal complex / mRNA branch site recognition / U2-type post-mRNA release spliceosomal complex / cellular bud site selection / pre-mRNA 3'-splice site binding / post-mRNA release spliceosomal complex / nuclear mRNA surveillance / generation of catalytic spliceosome for first transesterification step ...U2-type post-spliceosomal complex / spliceosomal complex disassembly / post-spliceosomal complex / mRNA branch site recognition / U2-type post-mRNA release spliceosomal complex / cellular bud site selection / pre-mRNA 3'-splice site binding / post-mRNA release spliceosomal complex / nuclear mRNA surveillance / generation of catalytic spliceosome for first transesterification step / cis assembly of pre-catalytic spliceosome / splicing factor binding / spliceosome conformational change to release U4 (or U4atac) and U1 (or U11) / U4/U6 snRNP / 7-methylguanosine cap hypermethylation / pICln-Sm protein complex / U2-type catalytic step 1 spliceosome / pre-mRNA binding / snRNP binding / small nuclear ribonucleoprotein complex / SMN-Sm protein complex / spliceosomal tri-snRNP complex / U2-type spliceosomal complex / commitment complex / mRNA cis splicing, via spliceosome / U2-type prespliceosome assembly / U2-type catalytic step 2 spliceosome / U4 snRNP / U2 snRNP / U1 snRNP / poly(U) RNA binding / U2-type prespliceosome / precatalytic spliceosome / generation of catalytic spliceosome for second transesterification step / Formation of TC-NER Pre-Incision Complex / spliceosomal complex assembly / mRNA 5'-splice site recognition / Gap-filling DNA repair synthesis and ligation in TC-NER / mRNA 3'-splice site recognition / DNA replication origin binding / Dual incision in TC-NER / spliceosomal tri-snRNP complex assembly / Prp19 complex / protein K63-linked ubiquitination / U5 snRNA binding / DNA replication initiation / U5 snRNP / U2 snRNA binding / U6 snRNA binding / pre-mRNA intronic binding / spliceosomal snRNP assembly / U1 snRNA binding / U4/U6 x U5 tri-snRNP complex / positive regulation of cell cycle / catalytic step 2 spliceosome / nuclear periphery / positive regulation of RNA splicing / spliceosomal complex / RING-type E3 ubiquitin transferase / mRNA splicing, via spliceosome / double-strand break repair via nonhomologous end joining / metallopeptidase activity / ubiquitin-protein transferase activity / ubiquitin protein ligase activity / chromosome, telomeric region / RNA helicase activity / RNA helicase / DNA repair / GTPase activity / mRNA binding / chromatin binding / GTP binding / chromatin / ATP hydrolysis activity / mitochondrion / DNA binding / RNA binding / zinc ion binding / ATP binding / identical protein binding / nucleus / cytosol / cytoplasm
Similarity search - Function
FAM192A/Fyv6, N-terminal / FAM192A/Fyv6, N-terminal domain / Prp18 / Pre-mRNA-splicing factor 18 / Prp18 domain / Pre-mRNA-splicing factor SLU7 domain / Pre-mRNA-splicing factor SLU7 / Pre-mRNA splicing Prp18-interacting factor / DHX8/ Prp22, DEXH-box helicase domain / : ...FAM192A/Fyv6, N-terminal / FAM192A/Fyv6, N-terminal domain / Prp18 / Pre-mRNA-splicing factor 18 / Prp18 domain / Pre-mRNA-splicing factor SLU7 domain / Pre-mRNA-splicing factor SLU7 / Pre-mRNA splicing Prp18-interacting factor / DHX8/ Prp22, DEXH-box helicase domain / : / : / Pre-mRNA-splicing factor Isy1 / Pre-mRNA-splicing factor Isy1 superfamily / Isy1-like splicing family / cwf21 / Slt11, RNA recognition motif / Pre-mRNA-splicing factor Cwc2, RNA recognition motif / Torus domain / Torus domain / mRNA splicing factor SYF2 / SYF2 splicing factor / RNA recognition motif. (a.k.a. RRM, RBD, or RNP domain) / Helix hairpin bin domain superfamily / : / : / mRNA splicing factor Cwf21 domain / cwf21 domain / Pre-mRNA-processing factor 17 / : / : / Pre-mRNA-splicing factor SYF1 middle HAT repeat / Pre-mRNA-splicing factor 19 / Pre-mRNA-processing factor 19 / Prp19/Pso4-like / G10 protein signature 2. / BUD31/G10-related, conserved site / G10 protein signature 1. / : / : / Pre-mRNA-splicing factor Syf1/CRNKL1 C-terminal HAT repeat / : / STL11, N-terminal / : / Pre-mRNA-splicing factor Syf1/CNRKL1 N-terminal HAT repeat / SKI-interacting protein SKIP, SNW domain / SKI-interacting protein, SKIP / SKIP/SNW domain / Pre-mRNA-splicing factor Cwf15/Cwc15 / Cwf15/Cwc15 cell cycle control protein / WD repeat Prp46/PLRG1-like / Pre-mRNA-splicing factor Cwc2/Slt11 / G10 protein / Pre-mRNA-splicing factor BUD31 / Pre-mRNA splicing factor component Cdc5p/Cef1, C-terminal / : / : / pre-mRNA splicing factor component / Initiation factor eIF-4 gamma, MA3 / MA3 domain / MI domain profile. / Domain in DAP-5, eIF4G, MA-3 and other proteins. / U2 small nuclear ribonucleoprotein A' / Middle domain of eukaryotic initiation factor 4G (eIF4G) / MIF4G-like, type 3 / U-box domain profile. / U2A'/phosphoprotein 32 family A, C-terminal / occurring C-terminal to leucine-rich repeats / Modified RING finger domain / U-box domain / Leucine-rich repeat / 116kDa U5 small nuclear ribonucleoprotein component, N-terminal / 116kDa U5 small nuclear ribonucleoprotein component, C-terminal / Snu114, GTP-binding domain / 116 kDa U5 small nuclear ribonucleoprotein component N-terminus / Pre-mRNA-splicing factor Syf1-like / : / Helicase associated domain (HA2), ratchet-like / DEAD-box helicase, OB fold / Oligonucleotide/oligosaccharide-binding (OB)-fold / Helicase associated domain (HA2), winged-helix / Helicase-associated domain / Helicase associated domain (HA2) Add an annotation / 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 / Sm-like protein Lsm6/SmF / Myb-type HTH DNA-binding domain profile. / Zinc finger, CCCH-type / Zinc finger C3H1-type profile. / LSM domain / LSM domain, eukaryotic/archaea-type / snRNP Sm proteins / HAT (Half-A-TPR) repeat / HAT (Half-A-TPR) repeats / Myb domain / :
Similarity search - Domain/homology
GUANOSINE-5'-TRIPHOSPHATE / INOSITOL HEXAKISPHOSPHATE / : / : / : / : / : / : / RNA / RNA (> 10) ...GUANOSINE-5'-TRIPHOSPHATE / INOSITOL HEXAKISPHOSPHATE / : / : / : / : / : / : / RNA / RNA (> 10) / RNA (> 100) / RNA (> 1000) / Pre-mRNA-splicing factor ISY1 / Pre-mRNA-splicing factor ATP-dependent RNA helicase PRP22 / Pre-mRNA-splicing factor BUD31 / Pre-mRNA-processing protein 45 / Pre-mRNA-processing factor 19 / Pre-mRNA-splicing factor 8 / Pre-mRNA-splicing factor 18 / Pre-mRNA-splicing factor SNU114 / Pre-mRNA-splicing factor SLT11 / Pre-mRNA-splicing factor NTC20 / Small nuclear ribonucleoprotein-associated protein B / Small nuclear ribonucleoprotein G / U2 small nuclear ribonucleoprotein B'' / Pre-mRNA-processing factor 17 / Small nuclear ribonucleoprotein Sm D3 / Pre-mRNA-splicing factor SYF2 / Pre-mRNA-splicing factor CWC22 / Protein FYV6 / Small nuclear ribonucleoprotein F / Small nuclear ribonucleoprotein Sm D1 / Pre-mRNA-splicing factor SLU7 / Pre-mRNA-splicing factor CWC21 / Pre-mRNA-splicing factor CEF1 / Pre-mRNA-splicing factor CWC15 / Pre-mRNA-splicing factor SYF1 / Pre-mRNA-splicing factor SNT309 / Small nuclear ribonucleoprotein Sm D2 / U2 small nuclear ribonucleoprotein A' / Pre-mRNA-splicing factor CWC2 / Pre-mRNA-splicing factor CLF1 / Small nuclear ribonucleoprotein E / Pre-mRNA-splicing factor PRP46
Similarity search - Component
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.31 Å
AuthorsWilkinson, M.E. / Hoskins, A.A.
Funding support United States, 1items
OrganizationGrant numberCountry
Howard Hughes Medical Institute (HHMI)HHMI Helen Hay Whitney Postdoctoral Fellowship United States
CitationJournal: bioRxiv / Year: 2024
Title: Control of 3' splice site selection by the yeast splicing factor Fyv6.
Authors: Katherine A Senn / Karli A Lipinski / Natalie J Zeps / Amory F Griffin / Max E Wilkinson / Aaron A Hoskins /
Abstract: Pre-mRNA splicing is catalyzed in two steps: 5' splice site (SS) cleavage and exon ligation. A number of proteins transiently associate with spliceosomes to specifically impact these steps (1 and 2 ...Pre-mRNA splicing is catalyzed in two steps: 5' splice site (SS) cleavage and exon ligation. A number of proteins transiently associate with spliceosomes to specifically impact these steps (1 and 2 step factors). We recently identified Fyv6 (FAM192A in humans) as a 2 step factor in ; however, we did not determine how widespread Fyv6's impact is on the transcriptome. To answer this question, we have used RNA-seq to analyze changes in splicing. These results show that loss of Fyv6 results in activation of non-consensus, branch point (BP) proximal 3' SS transcriptome-wide. To identify the molecular basis of these observations, we determined a high-resolution cryo-EM structure of a yeast product complex spliceosome containing Fyv6 at 2.3 Å. The structure reveals that Fyv6 is the only 2 step factor that contacts the Prp22 ATPase and that Fyv6 binding is mutually exclusive with that of the 1 step factor Yju2. We then use this structure to dissect Fyv6 functional domains and interpret results of a genetic screen for suppressor mutations. The combined transcriptomic, structural, and genetic studies allow us to propose a model in which Yju2/Fyv6 exchange facilitates exon ligation and Fyv6 promotes usage of consensus, BP distal 3' SS.
History
DepositionOct 1, 2024Deposition site: RCSB / Processing site: RCSB
Revision 1.0Dec 25, 2024Provider: repository / Type: Initial release
Revision 1.1Jan 1, 2025Group: Data collection / Database references / Category: citation / citation_author / em_admin
Item: _citation.journal_abbrev / _citation.journal_id_ISSN ..._citation.journal_abbrev / _citation.journal_id_ISSN / _citation.pdbx_database_id_DOI / _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
2: U2 snRNA
5: U5 snRNA
6: U6 snRNA
A: Pre-mRNA-splicing factor 8
C: Pre-mRNA-splicing factor SNU114
D: Protein FYV6
E: UBC4 mRNA spliced exons
G: Pre-mRNA-splicing factor ISY1
H: Pre-mRNA-splicing factor CWC22
I: UBC4 lariat-intron
J: Pre-mRNA-splicing factor PRP46
K: Pre-mRNA-processing protein 45
L: Pre-mRNA-splicing factor BUD31
M: Pre-mRNA-splicing factor CWC2
N: Pre-mRNA-splicing factor SLT11
O: Pre-mRNA-splicing factor CEF1
P: Pre-mRNA-splicing factor CWC15
R: Pre-mRNA-splicing factor CWC21
S: Pre-mRNA-splicing factor CLF1
T: Pre-mRNA-splicing factor SYF1
V: Pre-mRNA-splicing factor ATP-dependent RNA helicase PRP22
W: U2 small nuclear ribonucleoprotein A'
Y: U2 small nuclear ribonucleoprotein B''
Z: Pre-mRNA-splicing factor NTC20
a: Pre-mRNA-splicing factor 18
b: Small nuclear ribonucleoprotein-associated protein 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
j: Small nuclear ribonucleoprotein Sm D2
k: Small nuclear ribonucleoprotein-associated protein 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 SNT309
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
hetero molecules


Theoretical massNumber of molelcules
Total (without water)2,280,69263
Polymers2,278,18647
Non-polymers2,50616
Water181
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

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RNA chain , 5 types, 5 molecules 256EI

#1: RNA chain U2 snRNA


Mass: 376267.406 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: GenBank: NR_132154
#2: RNA chain U5 snRNA


Mass: 68643.344 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: GenBank: NR_132184
#3: RNA chain U6 snRNA


Mass: 35883.176 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: GenBank: NR_132206
#7: RNA chain UBC4 mRNA spliced exons


Mass: 13440.030 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Saccharomyces cerevisiae (brewer's yeast) / References: GenBank: NM_001178430
#10: RNA chain UBC4 lariat-intron


Mass: 30200.730 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Saccharomyces cerevisiae (brewer's yeast) / References: GenBank: 1931130107

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Pre-mRNA-splicing factor ... , 19 types, 19 molecules ACGHJLMNOPRSTVZacsy

#4: Protein Pre-mRNA-splicing factor 8


Mass: 279867.469 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P33334
#5: Protein Pre-mRNA-splicing factor SNU114 / 114 kDa U5 small nuclear ribonucleoprotein component / Growth inhibitory protein 10


Mass: 114174.008 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P36048
#8: Protein Pre-mRNA-splicing factor ISY1 / Interactor of SYF1 / PRP19-associated complex protein 30


Mass: 28073.836 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P21374
#9: Protein Pre-mRNA-splicing factor CWC22 / Complexed with CEF1 protein 22


Mass: 67386.062 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P53333
#11: Protein Pre-mRNA-splicing factor PRP46 / Complexed with CEF1 protein 1 / PRP nineteen-associated complex protein 50 / PRP19-associated ...Complexed with CEF1 protein 1 / PRP nineteen-associated complex protein 50 / PRP19-associated complex protein 50 / Pre-mRNA-processing protein 46


Mass: 50771.289 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: Q12417
#13: Protein Pre-mRNA-splicing factor BUD31 / Bud site selection protein 31 / Complexed with CEF1 protein 14


Mass: 18484.502 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P25337
#14: Protein Pre-mRNA-splicing factor CWC2 / Complexed with CEF1 protein 2 / PRP19-associated complex protein 40 / Synthetic lethal with CLF1 protein 3


Mass: 38486.562 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: Q12046
#15: Protein Pre-mRNA-splicing factor SLT11 / Extracellular mutant protein 2 / Synthetic lethality with U2 protein 11


Mass: 40988.590 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P38241
#16: Protein Pre-mRNA-splicing factor CEF1 / PRP nineteen-associated complex protein 85 / PRP19-associated complex protein 85


Mass: 67837.773 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: Q03654
#17: Protein Pre-mRNA-splicing factor CWC15 / Complexed with CEF1 protein 15


Mass: 19975.195 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: Q03772
#18: Protein Pre-mRNA-splicing factor CWC21 / Complexed with CEF1 protein 21


Mass: 15793.596 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: Q03375
#19: Protein Pre-mRNA-splicing factor CLF1 / Crooked neck-like factor 1 / PRP19-associated complex protein 77 / Synthetic lethal with CDC40 protein 3


Mass: 82555.859 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: Q12309
#20: Protein Pre-mRNA-splicing factor SYF1 / PRP19-associated complex protein 90 / Synthetic lethal with CDC40 protein 1


Mass: 100344.016 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: Q04048
#21: Protein Pre-mRNA-splicing factor ATP-dependent RNA helicase PRP22


Mass: 130171.359 Da / Num. of mol.: 1 / Mutation: S635A
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: PRP22, YER013W / Production host: Saccharomyces cerevisiae (brewer's yeast) / Strain (production host): BCY123 / References: UniProt: P24384, RNA helicase
#24: Protein Pre-mRNA-splicing factor NTC20 / PRP19-associated complex protein 20 / Synergistic to PRP19 mutation protein 384


Mass: 15992.927 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P38302
#25: Protein Pre-mRNA-splicing factor 18


Mass: 28414.391 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P33411
#27: Protein Pre-mRNA-splicing factor SLU7 / Synthetic lethal with U2 snRNA protein 17 / Synthetic lethal with U5 snRNA protein 7


Mass: 44722.875 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: Q02775
#35: Protein Pre-mRNA-splicing factor SNT309 / PRP19-associated complex protein 25 / Synergistic to PRP19 mutation protein 309


Mass: 20741.455 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: Q06091
#37: Protein Pre-mRNA-splicing factor SYF2 / PRP19 complex protein 31 / Synthetic lethal with CDC40 protein 2


Mass: 24850.719 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P53277

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Protein , 3 types, 4 molecules DKbk

#6: Protein Protein FYV6 / Function required for yeast viability protein 6


Mass: 20066.938 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P53913
#12: Protein Pre-mRNA-processing protein 45


Mass: 42548.727 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P28004
#26: Protein Small nuclear ribonucleoprotein-associated protein B / snRNP-B / Sm protein B / Sm-B / SmB


Mass: 22426.990 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P40018

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

#22: Protein U2 small nuclear ribonucleoprotein A' / U2 snRNP A' / Looks exceptionally like U2A protein 1


Mass: 27232.252 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: Q08963
#23: Protein U2 small nuclear ribonucleoprotein B'' / U2 snRNP B''


Mass: 12850.944 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P40567

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Small nuclear ribonucleoprotein ... , 6 types, 12 molecules dnepfqgrhljm

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


Mass: 11240.139 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P43321
#29: Protein Small nuclear ribonucleoprotein E / snRNP-E / Sm protein E / Sm-E / SmE


Mass: 10385.098 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: Q12330
#30: Protein Small nuclear ribonucleoprotein F / snRNP-F / Sm protein F / Sm-F / SmF


Mass: 9669.945 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P54999
#31: Protein Small nuclear ribonucleoprotein G / snRNP-G / Sm protein G / Sm-G / SmG


Mass: 8490.809 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P40204
#32: Protein Small nuclear ribonucleoprotein Sm D1 / Sm-D1 / snRNP core protein D1


Mass: 16296.798 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: Q02260
#33: Protein Small nuclear ribonucleoprotein Sm D2 / Sm-D2 / snRNP core protein D2


Mass: 12876.066 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: Q06217

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

#34: Protein Pre-mRNA-processing factor 17 / Cell division control protein 40


Mass: 52128.762 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P40968
#36: Protein
Pre-mRNA-processing factor 19 / RING-type E3 ubiquitin transferase PRP19


Mass: 56629.777 Da / Num. of mol.: 4 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast)
References: UniProt: P32523, RING-type E3 ubiquitin transferase

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Non-polymers , 6 types, 17 molecules

#38: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Mg
#39: Chemical
ChemComp-K / POTASSIUM ION


Mass: 39.098 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: K
#40: Chemical ChemComp-IHP / INOSITOL HEXAKISPHOSPHATE / MYO-INOSITOL HEXAKISPHOSPHATE / INOSITOL 1,2,3,4,5,6-HEXAKISPHOSPHATE


Mass: 660.035 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C6H18O24P6
#41: Chemical ChemComp-GTP / GUANOSINE-5'-TRIPHOSPHATE


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


Mass: 65.409 Da / Num. of mol.: 7 / Source method: obtained synthetically / Formula: Zn
#43: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Formula: H2O

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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: Yeast post-catalytic P complex spliceosome / Type: COMPLEX / Entity ID: #1-#37 / Source: NATURAL
Molecular weightValue: 2.27 MDa / Experimental value: NO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Buffer solutionpH: 7.9
Buffer component
IDConc.NameBuffer-ID
120 mMHEPES1
275 mMpotassium chloride1
30.25 mMEDTA1
SpecimenConc.: 0.1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R2/2
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal magnification: 130000 X / Nominal defocus max: 3100 nm / Nominal defocus min: 1300 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 0.72 sec. / Electron dose: 40 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 2 / Num. of real images: 51113

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Processing

EM software
IDNameVersionCategoryDetails
1Topazparticle selection
2EPUimage acquisition
4CTFFIND4.1CTF correction
7Coot0.9.8.3model fitting
9RELION5initial Euler assignment
10RELION5final Euler assignment
11RELION5classification
12RELION53D reconstructionBlush regularization used
13ISOLDE1.7model refinement
14PHENIXmodel refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 1820457
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 2.31 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 209005 / Num. of class averages: 1 / Symmetry type: POINT
Atomic model buildingProtocol: FLEXIBLE FIT / Space: REAL
Details: After fixing in ISOLDE, the model was refined in phenix.real_space_refine with the following non-default flags: refinement { run = *minimization_global rigid_body local_grid_search morphing ...Details: After fixing in ISOLDE, the model was refined in phenix.real_space_refine with the following non-default flags: refinement { run = *minimization_global rigid_body local_grid_search morphing simulated_annealing *adp } reference_model { enabled = True use_starting_model_as_reference = True fix_outliers = False strict_rotamer_matching = True } pdb_interpretation { reference_coordinate_restraints { enabled = True exclude_outliers = False top_out = True } automatic_linking { link_metals = True } peptide_link { restrain_rama_outliers = False restrain_rama_allowed = False } ramachandran_plot_restraints { enabled = False } }
Atomic model buildingPDB-ID: 6EXN

6exn


Accession code: 6EXN / Source name: PDB / Type: experimental model

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