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- EMDB-4225: Cryo-EM structure of the human CPSF160-WDR33-CPSF30 complex bound... -

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Basic information

Entry
Database: EMDB / ID: EMD-4225
TitleCryo-EM structure of the human CPSF160-WDR33-CPSF30 complex bound to the PAS AAUAAA motif at 3.1 Angstrom resolution.
Map dataOutput from Relion2 PostProcess
Sample
  • Complex: Human CPSF160-WDR33-CPSF30 heterotrimer bound to the AAUAAA motif of the Polyadenylation Signal
    • Complex: Human CPSF160-WDR33-CPSF30 heterotrimer
      • Protein or peptide: Cleavage and polyadenylation specificity factor subunit 1
      • Protein or peptide: pre-mRNA 3' end processing protein WDR33
      • Protein or peptide: Cleavage and polyadenylation specificity factor subunit 4
    • Complex: AAUAAA motif of the Polyadenylation Signal
      • RNA: RNA (5'-R(P*AP*AP*UP*AP*AP*AP*GP*G)-3')
  • Ligand: ZINC ION
Function / homology
Function and homology information


co-transcriptional RNA 3'-end processing, cleavage and polyadenylation pathway / Inhibition of Host mRNA Processing and RNA Silencing / Processing of Intronless Pre-mRNAs / mRNA cleavage and polyadenylation specificity factor complex / mRNA 3'-UTR AU-rich region binding / collagen trimer / mRNA 3'-end processing / Transport of Mature mRNA Derived from an Intronless Transcript / postreplication repair / tRNA processing in the nucleus ...co-transcriptional RNA 3'-end processing, cleavage and polyadenylation pathway / Inhibition of Host mRNA Processing and RNA Silencing / Processing of Intronless Pre-mRNAs / mRNA cleavage and polyadenylation specificity factor complex / mRNA 3'-UTR AU-rich region binding / collagen trimer / mRNA 3'-end processing / Transport of Mature mRNA Derived from an Intronless Transcript / postreplication repair / tRNA processing in the nucleus / RNA Polymerase II Transcription Termination / : / Processing of Capped Intron-Containing Pre-mRNA / mRNA processing / fibrillar center / sequence-specific double-stranded DNA binding / spermatogenesis / intracellular membrane-bounded organelle / enzyme binding / RNA binding / zinc ion binding / nucleoplasm / nucleus
Similarity search - Function
Zinc-finger CCCH domain / Zinc-finger containing family / CPSF complex subunit CPSF4-like / Pre-mRNA 3' end processing protein Pfs2-like / Zinc finger C-x8-C-x5-C-x3-H type (and similar) / Zinc finger, CCCH-type superfamily / zinc finger / Zinc finger, CCCH-type / Zinc finger C3H1-type profile. / Collagen triple helix repeat ...Zinc-finger CCCH domain / Zinc-finger containing family / CPSF complex subunit CPSF4-like / Pre-mRNA 3' end processing protein Pfs2-like / Zinc finger C-x8-C-x5-C-x3-H type (and similar) / Zinc finger, CCCH-type superfamily / zinc finger / Zinc finger, CCCH-type / Zinc finger C3H1-type profile. / Collagen triple helix repeat / Collagen triple helix repeat (20 copies) / 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 / zinc finger / Zinc knuckle / Zinc finger, CCHC-type superfamily / Zinc finger, CCHC-type / Zinc finger CCHC-type profile. / WD domain, G-beta repeat / WD40 repeats / WD40 repeat / Trp-Asp (WD) repeats profile. / Trp-Asp (WD) repeats circular profile. / WD40-repeat-containing domain superfamily / WD40/YVTN repeat-like-containing domain superfamily
Similarity search - Domain/homology
Cleavage and polyadenylation specificity factor subunit 4 / Cleavage and polyadenylation specificity factor subunit 1 / pre-mRNA 3' end processing protein WDR33
Similarity search - Component
Biological speciesHomo sapiens (human) / unidentified adenovirus
Methodsingle particle reconstruction / cryo EM / Resolution: 3.07 Å
AuthorsClerici M / Faini M / Jinek M
Funding support Belgium, Germany, 4 items
OrganizationGrant numberCountry
European Research CouncilERC-StG-337284 Belgium
European Molecular Biology OrganizationEMBO ALTF-343-2013 Germany
European Research CouncilERC Advanced Grants no. 233226 and no. 670821 Belgium
European UnionPROSPECTS, HEALTH-F4-2008-201648 Belgium
CitationJournal: Nat Struct Mol Biol / Year: 2018
Title: Structural basis of AAUAAA polyadenylation signal recognition by the human CPSF complex.
Authors: Marcello Clerici / Marco Faini / Lena M Muckenfuss / Ruedi Aebersold / Martin Jinek /
Abstract: Mammalian mRNA biogenesis requires specific recognition of a hexanucleotide AAUAAA motif in the polyadenylation signals (PAS) of precursor mRNA (pre-mRNA) transcripts by the cleavage and ...Mammalian mRNA biogenesis requires specific recognition of a hexanucleotide AAUAAA motif in the polyadenylation signals (PAS) of precursor mRNA (pre-mRNA) transcripts by the cleavage and polyadenylation specificity factor (CPSF) complex. Here we present a 3.1-Å-resolution cryo-EM structure of a core CPSF module bound to the PAS hexamer motif. The structure reveals the molecular interactions responsible for base-specific recognition, providing a rationale for mechanistic differences between mammalian and yeast 3' polyadenylation.
History
DepositionDec 19, 2017-
Header (metadata) releaseJan 3, 2018-
Map releaseJan 31, 2018-
UpdateMar 1, 2023-
Current statusMar 1, 2023Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.058
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.058
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6fuw
  • Surface level: 0.058
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_4225.png

Downloads & links

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Map

FileDownload / File: emd_4225.map.gz / Format: CCP4 / Size: 30.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationOutput from Relion2 PostProcess
Voxel sizeX=Y=Z: 1.058 Å
Density
Contour LevelBy AUTHOR: 0.058 / Movie #1: 0.058
Minimum - Maximum-0.2077625 - 0.32876167
Average (Standard dev.)0.00030825817 (±0.0140046645)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions200200200
Spacing200200200
CellA=B=C: 211.59999 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.0581.0581.058
M x/y/z200200200
origin x/y/z0.0000.0000.000
length x/y/z211.600211.600211.600
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ240240240
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS200200200
D min/max/mean-0.2080.3290.000

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Supplemental data

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Mask #1

Fileemd_4225_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Human CPSF160-WDR33-CPSF30 heterotrimer bound to the AAUAAA motif...

EntireName: Human CPSF160-WDR33-CPSF30 heterotrimer bound to the AAUAAA motif of the Polyadenylation Signal
Components
  • Complex: Human CPSF160-WDR33-CPSF30 heterotrimer bound to the AAUAAA motif of the Polyadenylation Signal
    • Complex: Human CPSF160-WDR33-CPSF30 heterotrimer
      • Protein or peptide: Cleavage and polyadenylation specificity factor subunit 1
      • Protein or peptide: pre-mRNA 3' end processing protein WDR33
      • Protein or peptide: Cleavage and polyadenylation specificity factor subunit 4
    • Complex: AAUAAA motif of the Polyadenylation Signal
      • RNA: RNA (5'-R(P*AP*AP*UP*AP*AP*AP*GP*G)-3')
  • Ligand: ZINC ION

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Supramolecule #1: Human CPSF160-WDR33-CPSF30 heterotrimer bound to the AAUAAA motif...

SupramoleculeName: Human CPSF160-WDR33-CPSF30 heterotrimer bound to the AAUAAA motif of the Polyadenylation Signal
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#4
Molecular weightTheoretical: 220 KDa

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Supramolecule #2: Human CPSF160-WDR33-CPSF30 heterotrimer

SupramoleculeName: Human CPSF160-WDR33-CPSF30 heterotrimer / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1-#3
Source (natural)Organism: Homo sapiens (human)

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Supramolecule #3: AAUAAA motif of the Polyadenylation Signal

SupramoleculeName: AAUAAA motif of the Polyadenylation Signal / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #4
Source (natural)Organism: unidentified adenovirus

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Macromolecule #1: Cleavage and polyadenylation specificity factor subunit 1

MacromoleculeName: Cleavage and polyadenylation specificity factor subunit 1
type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 161.346484 KDa
Recombinant expressionOrganism: Spodoptera aff. frugiperda 2 RZ-2014 (butterflies/moths)
SequenceString: SNAMYAVYKQ AHPPTGLEFS MYCNFFNNSE RNLVVAGTSQ LYVYRLNRDA EALTKNDRST EGKAHREKLE LAASFSFFGN VMSMASVQL AGAKRDALLL SFKDAKLSVV EYDPGTHDLK TLSLHYFEEP ELRDGFVQNV HTPRVRVDPD GRCAAMLVYG T RLVVLPFR ...String:
SNAMYAVYKQ AHPPTGLEFS MYCNFFNNSE RNLVVAGTSQ LYVYRLNRDA EALTKNDRST EGKAHREKLE LAASFSFFGN VMSMASVQL AGAKRDALLL SFKDAKLSVV EYDPGTHDLK TLSLHYFEEP ELRDGFVQNV HTPRVRVDPD GRCAAMLVYG T RLVVLPFR RESLAEEHEG LVGEGQRSSF LPSYIIDVRA LDEKLLNIID LQFLHGYYEP TLLILFEPNQ TWPGRVAVRQ DT CSIVAIS LNITQKVHPV IWSLTSLPFD CTQALAVPKP IGGVVVFAVN SLLYLNQSVP PYGVALNSLT TGTTAFPLRT QEG VRITLD CAQATFISYD KMVISLKGGE IYVLTLITDG MRSVRAFHFD KAAASVLTTS MVTMEPGYLF LGSRLGNSLL LKYT EKLQE PPASAVREAA DKEEPPSKKK RVDATAGWSA AGKSVPQDEV DEIEVYGSEA QSGTQLATYS FEVCDSILNI GPCAN AAVG EPAFLSEEFQ NSPEPDLEIV VCSGHGKNGA LSVLQKSIRP QVVTTFELPG CYDMWTVIAP VRKEEEDNPK GEGTEQ EPS TTPEADDDGR RHGFLILSRE DSTMILQTGQ EIMELDTSGF ATQGPTVFAG NIGDNRYIVQ VSPLGIRLLE GVNQLHF IP VDLGAPIVQC AVADPYVVIM SAEGHVTMFL LKSDSYGGRH HRLALHKPPL HHQSKVITLC LYRDLSGMFT TESRLGGA R DELGGRSGPE AEGLGSETSP TVDDEEEMLY GDSGSLFSPS KEEARRSSQP PADRDPAPFR AEPTHWCLLV RENGTMEIY QLPDWRLVFL VKNFPVGQRV LVDSSFGQPT TQGEARREEA TRQGELPLVK EVLLVALGSR QSRPYLLVHV DQELLIYEAF PHDSQLGQG NLKVRFKKVP HNINFREKKP KPSKKKAEGG GAEEGAGARG RVARFRYFED IYGYSGVFIC GPSPHWLLVT G RGALRLHP MAIDGPVDSF APFHNVNCPR GFLYFNRQGE LRISVLPAYL SYDAPWPVRK IPLRCTAHYV AYHVESKVYA VA TSTNTPC ARIPRMTGEE KEFETIERDE RYIHPQQEAF SIQLISPVSW EAIPNARIEL QEWEHVTCMK TVSLRSEETV SGL KGYVAA GTCLMQGEEV TCRGRILIMD VIEVVPEPGQ PLTKNKFKVL YEKEQKGPVT ALCHCNGHLV SAIGQKIFLW SLRA SELTG MAFIDTQLYI HQMISVKNFI LAADVMKSIS LLRYQEESKT LSLVSRDAKP LEVYSVDFMV DNAQLGFLVS DRDRN LMVY MYLPEAKESF GGMRLLRRAD FHVGAHVNTF WRTPCRGATE GLSKKSVVWE NKHITWFATL DGGIGLLLPM QEKTYR RLL MLQNALTTML PHHAGLNPRA FRMLHVDRRT LQNAVRNVLD GELLNRYLYL STMERSELAK KIGTTPDIIL DDLLETD RV TAHF

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Macromolecule #2: pre-mRNA 3' end processing protein WDR33

MacromoleculeName: pre-mRNA 3' end processing protein WDR33 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 47.44891 KDa
Recombinant expressionOrganism: Spodoptera aff. frugiperda 2 RZ-2014 (butterflies/moths)
SequenceString: SNAMATEIGS PPRFFHMPRF QHQAPRQLFY KRPDFAQQQA MQQLTFDGKR MRKAVNRKTI DYNPSVIKYL ENRIWQRDQR DMRAIQPDA GYYNDLVPPI GMLNNPMNAV TTKFVRTSTN KVKCPVFVVR WTPEGRRLVT GASSGEFTLW NGLTFNFETI L QAHDSPVR ...String:
SNAMATEIGS PPRFFHMPRF QHQAPRQLFY KRPDFAQQQA MQQLTFDGKR MRKAVNRKTI DYNPSVIKYL ENRIWQRDQR DMRAIQPDA GYYNDLVPPI GMLNNPMNAV TTKFVRTSTN KVKCPVFVVR WTPEGRRLVT GASSGEFTLW NGLTFNFETI L QAHDSPVR AMTWSHNDMW MLTADHGGYV KYWQSNMNNV KMFQAHKEAI REASFSPTDN KFATCSDDGT VRIWDFLRCH EE RILRGHG ADVKCVDWHP TKGLVVSGSK DSQQPIKFWD PKTGQSLATL HAHKNTVMEV KLNLNGNWLL TASRDHLCKL FDI RNLKEE LQVFRGHKKE ATAVAWHPVH EGLFASGGSD GSLLFWHVGV EKEVGGMEMA HEGMIWSLAW HPLGHILCSG SNDH TSKFW TRNRPGDK

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Macromolecule #3: Cleavage and polyadenylation specificity factor subunit 4

MacromoleculeName: Cleavage and polyadenylation specificity factor subunit 4
type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 20.422855 KDa
Recombinant expressionOrganism: Spodoptera aff. frugiperda 2 RZ-2014 (butterflies/moths)
SequenceString:
MQEIIASVDH IKFDLEIAVE QQLGAQPLPF PGMDKSGAAV CEFFLKAACG KGGMCPFRHI SGEKTVVCKH WLRGLCKKGD QCEFLHEYD MTKMPECYFY SKFGECSNKE CPFLHIDPES KIKDCPWYDR GFCKHGPLCR HRHTRRVICV NYLVGFCPEG P SCKFMHPR FELPMGTTEQ

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Macromolecule #4: RNA (5'-R(P*AP*AP*UP*AP*AP*AP*GP*G)-3')

MacromoleculeName: RNA (5'-R(P*AP*AP*UP*AP*AP*AP*GP*G)-3') / type: rna / ID: 4 / Number of copies: 1
Source (natural)Organism: unidentified adenovirus
Molecular weightTheoretical: 3.232036 KDa
SequenceString:
ACAAUAAAGG

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Macromolecule #5: ZINC ION

MacromoleculeName: ZINC ION / type: ligand / ID: 5 / Number of copies: 3 / Formula: ZN
Molecular weightTheoretical: 65.409 Da

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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

Concentration0.3 mg/mL
BufferpH: 7.5
Component:
ConcentrationFormulaName
140.0 mMKClPotassium Chloride
20.0 mMHEPESHEPES
1.0 mMDTTDithiothreitol
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec.
VitrificationCryogen name: ETHANE-PROPANE / Chamber humidity: 100 % / Chamber temperature: 293 K / Instrument: FEI VITROBOT MARK IV / Details: 15 seconds wait time prior to blotting.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 47259 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.2 µm / Nominal defocus min: 1.1 µm
Specialist opticsEnergy filter - Name: GIF Quantum LS
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Number grids imaged: 1 / Number real images: 1070 / Average exposure time: 10.0 sec. / Average electron dose: 80.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 263000
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

6f9n


Details: CPSF160-WDR33 heterodimer
Initial angle assignmentType: RANDOM ASSIGNMENT / Software - Name: RELION (ver. 2.1b1_cu8.0)
Final angle assignmentType: PROJECTION MATCHING / Software - Name: RELION (ver. 2.1b1_cu8.0)
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.07 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 2.1b1_cu8.0) / Number images used: 137000
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

6f9n


Details: fitted manually in Coot
Detailsphenix.real_space_refine
RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-6fuw:
Cryo-EM structure of the human CPSF160-WDR33-CPSF30 complex bound to the PAS AAUAAA motif at 3.1 Angstrom resolution

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