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- EMDB-6684: Cryo-EM structure of the Catalytic Step II spliceosome (C* comple... -

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Basic information

Entry
Database: EMDB / ID: EMD-6684
TitleCryo-EM structure of the Catalytic Step II spliceosome (C* complex) at 4.0 angstrom resolution
Map dataYeast Step II Catalytically Activated Spliceosome, C* spliceosome
Sample
  • Complex: C* spliceosome
Function / homology
Function and homology information


RNA exon ligation / snRNA metabolic process / snRNA modification / U2-type post-spliceosomal complex / mRNA branch site recognition / U2-type post-mRNA release spliceosomal complex / cellular bud site selection / post-mRNA release spliceosomal complex / 3'-5' RNA helicase activity / nuclear mRNA surveillance ...RNA exon ligation / snRNA metabolic process / snRNA modification / U2-type post-spliceosomal complex / mRNA branch site recognition / U2-type post-mRNA release spliceosomal complex / cellular bud site selection / post-mRNA release spliceosomal complex / 3'-5' RNA helicase activity / 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 / ATP-dependent activity, acting on RNA / 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 / metallopeptidase activity / ubiquitin-protein transferase activity / ubiquitin protein ligase 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
Prp18 / Pre-mRNA-splicing factor 18 / Prp18 domain / cwf21 / Slt11, RNA recognition motif / Pre-mRNA-splicing factor Cwc2, RNA recognition motif / Torus domain / Torus domain / mRNA splicing factor SYF2 / SYF2 splicing factor ...Prp18 / Pre-mRNA-splicing factor 18 / Prp18 domain / 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) / : / : / 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 / : / Myb-like DNA-binding domain / Sm domain profile. / DNA/RNA helicase, ATP-dependent, DEAH-box type, conserved site / DEAH-box subfamily ATP-dependent helicases signature. / LSM domain superfamily / Translation elongation factor EFG/EF2, domain IV / Elongation factor G, domain IV / Elongation factor G, domain IV / Elongation factor G C-terminus / Quinoprotein alcohol dehydrogenase-like superfamily / Elongation factor EFG, domain V-like / Elongation factor G C-terminus
Similarity search - Domain/homology
Pre-mRNA-splicing factor ATP-dependent RNA helicase PRP16 / 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 / Small nuclear ribonucleoprotein-associated protein B / Small nuclear ribonucleoprotein G ...Pre-mRNA-splicing factor ATP-dependent RNA helicase PRP16 / 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 / 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 / Small nuclear ribonucleoprotein F / Small nuclear ribonucleoprotein Sm D1 / 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 S288c (yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.0 Å
AuthorsYan C / Wan R / Bai R / Huang G / Shi Y
CitationJournal: Science / Year: 2017
Title: Structure of a yeast step II catalytically activated spliceosome.
Authors: Chuangye Yan / Ruixue Wan / Rui Bai / Gaoxingyu Huang / Yigong Shi /
Abstract: Each cycle of precursor messenger RNA (pre-mRNA) splicing comprises two sequential reactions, first freeing the 5' exon and generating an intron lariat-3' exon and then ligating the two exons and ...Each cycle of precursor messenger RNA (pre-mRNA) splicing comprises two sequential reactions, first freeing the 5' exon and generating an intron lariat-3' exon and then ligating the two exons and releasing the intron lariat. The second reaction is executed by the step II catalytically activated spliceosome (known as the C* complex). Here, we present the cryo-electron microscopy structure of a C* complex from Saccharomyces cerevisiae at an average resolution of 4.0 angstroms. Compared with the preceding spliceosomal complex (C complex), the lariat junction has been translocated by 15 to 20 angstroms to vacate space for the incoming 3'-exon sequences. The step I splicing factors Cwc25 and Yju2 have been dissociated from the active site. Two catalytic motifs from Prp8 (the 1585 loop and the β finger of the ribonuclease H-like domain), along with the step II splicing factors Prp17 and Prp18 and other surrounding proteins, are poised to assist the second transesterification. These structural features, together with those reported for other spliceosomal complexes, yield a near-complete mechanistic picture on the splicing cycle.
History
DepositionDec 6, 2016-
Header (metadata) releaseJan 25, 2017-
Map releaseJan 25, 2017-
UpdateJan 22, 2020-
Current statusJan 22, 2020Processing site: PDBj / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0297
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.0297
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-5wsg
  • Surface level: 0.0297
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_6684.png

Downloads & links

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Map

FileDownload / File: emd_6684.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationYeast Step II Catalytically Activated Spliceosome, C* spliceosome
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.31 Å/pix.
x 400 pix.
= 522.4 Å		1.31 Å/pix.
x 400 pix.
= 522.4 Å		1.31 Å/pix.
x 400 pix.
= 522.4 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.306 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0297 / Movie #1: 0.0297
Minimum - Maximum-0.09387038 - 0.19135746
Average (Standard dev.)-0.00000181478 (±0.006615814)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions400400400
Spacing400400400
CellA=B=C: 522.4 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.3061.3061.306
M x/y/z400400400
origin x/y/z0.0000.0000.000
length x/y/z522.400522.400522.400
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS400400400
D min/max/mean-0.0940.191-0.000

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Supplemental data

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Sample components

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Entire : C* spliceosome

EntireName: C* spliceosome
Components
  • Complex: C* spliceosome

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Supramolecule #1: C* spliceosome

SupramoleculeName: C* spliceosome / type: complex / ID: 1 / Parent: 0
Source (natural)Organism: Saccharomyces cerevisiae S288c (yeast)
Molecular weightTheoretical: 1.5 MDa

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.27 mg/mL
BufferpH: 8
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 1.6 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm
Sample stageCooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionSoftware - details: CTFFIND3
Startup modelType of model: EMDB MAP
EMDB ID:

6413

Final reconstructionResolution.type: BY AUTHOR / Resolution: 4.0 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 27558
Initial angle assignmentType: ANGULAR RECONSTITUTION
Final angle assignmentType: ANGULAR RECONSTITUTION

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