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- EMDB-20861: Cryo-EM structure of human CPSF160-WDR33-CPSF30-PAS RNA-CstF77 complex -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-20861 | |||||||||
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Title | Cryo-EM structure of human CPSF160-WDR33-CPSF30-PAS RNA-CstF77 complex | |||||||||
![]() | human CPSF160-WDR33-CPSF30-PAS RNA-CstF77 complex | |||||||||
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![]() | pre-mRNA 3'-end processing / RNA binding / AAUAAA polyadenylation signal / RNA BINDING PROTEIN-RNA complex / mPSF | |||||||||
Function / homology | ![]() mRNA cleavage stimulating factor complex / co-transcriptional RNA 3'-end processing, cleavage and polyadenylation pathway / Inhibition of Host mRNA Processing and RNA Silencing / RNA 3'-end processing / co-transcriptional mRNA 3'-end processing, cleavage and polyadenylation pathway / Processing of Intronless Pre-mRNAs / mRNA cleavage and polyadenylation specificity factor complex / collagen trimer / mRNA 3'-UTR AU-rich region binding / Transport of Mature mRNA Derived from an Intronless Transcript ...mRNA cleavage stimulating factor complex / co-transcriptional RNA 3'-end processing, cleavage and polyadenylation pathway / Inhibition of Host mRNA Processing and RNA Silencing / RNA 3'-end processing / co-transcriptional mRNA 3'-end processing, cleavage and polyadenylation pathway / Processing of Intronless Pre-mRNAs / mRNA cleavage and polyadenylation specificity factor complex / collagen trimer / mRNA 3'-UTR AU-rich region binding / Transport of Mature mRNA Derived from an Intronless Transcript / mRNA 3'-end processing / mRNA 3'-end processing / postreplication repair / tRNA processing in the nucleus / RNA Polymerase II Transcription Termination / Processing of Capped Intron-Containing Pre-mRNA / fibrillar center / mRNA processing / sequence-specific double-stranded DNA binding / spermatogenesis / intracellular membrane-bounded organelle / mRNA binding / enzyme binding / RNA binding / zinc ion binding / nucleoplasm / nucleus Similarity search - Function | |||||||||
Biological species | ![]() ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.6 Å | |||||||||
![]() | Sun Y / Zhang Y / Walz T / Tong L | |||||||||
Funding support | ![]()
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![]() | ![]() Title: Structural Insights into the Human Pre-mRNA 3'-End Processing Machinery. Authors: Yixiao Zhang / Yadong Sun / Yongsheng Shi / Thomas Walz / Liang Tong / ![]() Abstract: The mammalian pre-mRNA 3'-end-processing machinery consists of cleavage and polyadenylation specificity factor (CPSF), cleavage stimulation factor (CstF), and other proteins, but the overall ...The mammalian pre-mRNA 3'-end-processing machinery consists of cleavage and polyadenylation specificity factor (CPSF), cleavage stimulation factor (CstF), and other proteins, but the overall architecture of this machinery remains unclear. CPSF contains two functionally distinct modules: a cleavage factor (mCF) and a polyadenylation specificity factor (mPSF). Here, we have produced recombinant human CPSF and CstF and examined these factors by electron microscopy (EM). We find that mPSF is the organizational core of the machinery, while the conformations of mCF and CstF and the position of mCF relative to mPSF are highly variable. We have identified by cryo-EM a segment in CPSF100 that tethers mCF to mPSF, and we have named it the PSF interaction motif (PIM). Mutations in the PIM can abolish CPSF formation, indicating that it is a crucial contact in CPSF. We have also obtained reconstructions of mCF and CstF77 by cryo-EM, assembled around the mPSF core. | |||||||||
History |
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Structure visualization
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 78.6 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 20.3 KB 20.3 KB | Display Display | ![]() |
Images | ![]() | 140.6 KB | ||
Filedesc metadata | ![]() | 8.2 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Validation report
Summary document | ![]() | 573.3 KB | Display | ![]() |
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Full document | ![]() | 572.9 KB | Display | |
Data in XML | ![]() | 6.2 KB | Display | |
Data in CIF | ![]() | 7.1 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 6uroMC ![]() 6urgC C: citing same article ( M: atomic model generated by this map |
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Similar structure data |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Annotation | human CPSF160-WDR33-CPSF30-PAS RNA-CstF77 complex | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.07 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
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Sample components
-Entire : The complex of human mPSF-PAS RNA-CstF
Entire | Name: The complex of human mPSF-PAS RNA-CstF |
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Components |
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-Supramolecule #1: The complex of human mPSF-PAS RNA-CstF
Supramolecule | Name: The complex of human mPSF-PAS RNA-CstF / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#5 |
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Source (natural) | Organism: ![]() |
-Macromolecule #1: Cleavage and polyadenylation specificity factor subunit 1
Macromolecule | Name: Cleavage and polyadenylation specificity factor subunit 1 type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 161.074234 KDa |
Recombinant expression | Organism: ![]() |
Sequence | String: MYAVYKQAHP PTGLEFSMYC NFFNNSERNL VVAGTSQLYV YRLNRDAEAL TKNDRSTEGK AHREKLELAA SFSFFGNVMS MASVQLAGA KRDALLLSFK DAKLSVVEYD PGTHDLKTLS LHYFEEPELR DGFVQNVHTP RVRVDPDGRC AAMLVYGTRL V VLPFRRES ...String: MYAVYKQAHP PTGLEFSMYC NFFNNSERNL VVAGTSQLYV YRLNRDAEAL TKNDRSTEGK AHREKLELAA SFSFFGNVMS MASVQLAGA KRDALLLSFK DAKLSVVEYD PGTHDLKTLS LHYFEEPELR DGFVQNVHTP RVRVDPDGRC AAMLVYGTRL V VLPFRRES LAEEHEGLVG EGQRSSFLPS YIIDVRALDE KLLNIIDLQF LHGYYEPTLL ILFEPNQTWP GRVAVRQDTC SI VAISLNI TQKVHPVIWS LTSLPFDCTQ ALAVPKPIGG VVVFAVNSLL YLNQSVPPYG VALNSLTTGT TAFPLRTQEG VRI TLDCAQ ATFISYDKMV ISLKGGEIYV LTLITDGMRS VRAFHFDKAA ASVLTTSMVT MEPGYLFLGS RLGNSLLLKY TEKL QEPPA SAVREAADKE EPPSKKKRVD ATAGWSAAGK SVPQDEVDEI EVYGSEAQSG TQLATYSFEV CDSILNIGPC ANAAV GEPA FLSEEFQNSP EPDLEIVVCS GHGKNGALSV LQKSIRPQVV TTFELPGCYD MWTVIAPVRK EEEDNPKGEG TEQEPS TTP EADDDGRRHG FLILSREDST MILQTGQEIM ELDTSGFATQ GPTVFAGNIG DNRYIVQVSP LGIRLLEGVN QLHFIPV DL GAPIVQCAVA DPYVVIMSAE GHVTMFLLKS DSYGGRHHRL ALHKPPLHHQ SKVITLCLYR DLSGMFTTES RLGGARDE L GGRSGPEAEG LGSETSPTVD DEEEMLYGDS GSLFSPSKEE ARRSSQPPAD RDPAPFRAEP THWCLLVREN GTMEIYQLP DWRLVFLVKN FPVGQRVLVD SSFGQPTTQG EARREEATRQ GELPLVKEVL LVALGSRQSR PYLLVHVDQE LLIYEAFPHD SQLGQGNLK VRFKKVPHNI NFREKKPKPS KKKAEGGGAE EGAGARGRVA RFRYFEDIYG YSGVFICGPS PHWLLVTGRG A LRLHPMAI DGPVDSFAPF HNVNCPRGFL YFNRQGELRI SVLPAYLSYD APWPVRKIPL RCTAHYVAYH VESKVYAVAT ST NTPCARI PRMTGEEKEF ETIERDERYI HPQQEAFSIQ LISPVSWEAI PNARIELQEW EHVTCMKTVS LRSEETVSGL KGY VAAGTC LMQGEEVTCR GRILIMDVIE VVPEPGQPLT KNKFKVLYEK EQKGPVTALC HCNGHLVSAI GQKIFLWSLR ASEL TGMAF IDTQLYIHQM ISVKNFILAA DVMKSISLLR YQEESKTLSL VSRDAKPLEV YSVDFMVDNA QLGFLVSDRD RNLMV YMYL PEAKESFGGM RLLRRADFHV GAHVNTFWRT PCRGATEGLS KKSVVWENKH ITWFATLDGG IGLLLPMQEK TYRRLL MLQ NALTTMLPHH AGLNPRAFRM LHVDRRTLQN AVRNVLDGEL LNRYLYLSTM ERSELAKKIG TTPDIILDDL LETDRVT AH F UniProtKB: Cleavage and polyadenylation specificity factor subunit 1 |
-Macromolecule #2: pre-mRNA 3' end processing protein WDR33
Macromolecule | Name: pre-mRNA 3' end processing protein WDR33 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 67.546812 KDa |
Recombinant expression | Organism: ![]() |
Sequence | String: MGSSHHHHHH SSGLVMATEI GSPPRFFHMP RFQHQAPRQL FYKRPDFAQQ QAMQQLTFDG KRMRKAVNRK TIDYNPSVIK YLENRIWQR DQRDMRAIQP DAGYYNDLVP PIGMLNNPMN AVTTKFVRTS TNKVKCPVFV VRWTPEGRRL VTGASSGEFT L WNGLTFNF ...String: MGSSHHHHHH SSGLVMATEI GSPPRFFHMP RFQHQAPRQL FYKRPDFAQQ QAMQQLTFDG KRMRKAVNRK TIDYNPSVIK YLENRIWQR DQRDMRAIQP DAGYYNDLVP PIGMLNNPMN AVTTKFVRTS TNKVKCPVFV VRWTPEGRRL VTGASSGEFT L WNGLTFNF ETILQAHDSP VRAMTWSHND MWMLTADHGG YVKYWQSNMN NVKMFQAHKE AIREASFSPT DNKFATCSDD GT VRIWDFL RCHEERILRG HGADVKCVDW HPTKGLVVSG SKDSQQPIKF WDPKTGQSLA TLHAHKNTVM EVKLNLNGNW LLT ASRDHL CKLFDIRNLK EELQVFRGHK KEATAVAWHP VHEGLFASGG SDGSLLFWHV GVEKEVGGME MAHEGMIWSL AWHP LGHIL CSGSNDHTSK FWTRNRPGDK MRDRYNLNLL PGMSEDGVEY DDLEPNSLAV IPGMGIPEQL KLAMEQEQMG KDESN EIEM TIPGLDWGME EVMQKDQKKV PQKKVPYAKP IPAQFQQAWM QNKVPIPAPN EVLNDRKEDI KLEEKKKTQA EIEQEM ATL QYTNPQLLEQ LKIERLAQKQ VEQI UniProtKB: pre-mRNA 3' end processing protein WDR33 |
-Macromolecule #3: Cleavage and polyadenylation specificity factor subunit 4
Macromolecule | Name: Cleavage and polyadenylation specificity factor subunit 4 type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 28.417883 KDa |
Recombinant expression | Organism: ![]() |
Sequence | String: MQEIIASVDH IKFDLEIAVE QQLGAQPLPF PGMDKSGAAV CEFFLKAACG KGGMCPFRHI SGEKTVVCKH WLRGLCKKGD QCEFLHEYD MTKMPECYFY SKFGECSNKE CPFLHIDPES KIKDCPWYDR GFCKHGPLCR HRHTRRVICV NYLVGFCPEG P SCKFMHPR ...String: MQEIIASVDH IKFDLEIAVE QQLGAQPLPF PGMDKSGAAV CEFFLKAACG KGGMCPFRHI SGEKTVVCKH WLRGLCKKGD QCEFLHEYD MTKMPECYFY SKFGECSNKE CPFLHIDPES KIKDCPWYDR GFCKHGPLCR HRHTRRVICV NYLVGFCPEG P SCKFMHPR FELPMGTTEQ PPLPQQTQPP AKQRTPQVIG VMQSQNSSAG NRGPRPLEQV TCYKCGEKGH YANRCTKGHL AF LSGQHHH HHH UniProtKB: Cleavage and polyadenylation specificity factor subunit 4 |
-Macromolecule #5: Cleavage stimulation factor subunit 3
Macromolecule | Name: Cleavage stimulation factor subunit 3 / type: protein_or_peptide / ID: 5 / Number of copies: 2 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 83.039625 KDa |
Recombinant expression | Organism: ![]() |
Sequence | String: MSGDGATEQA AEYVPEKVKK AEKKLEENPY DLDAWSILIR EAQNQPIDKA RKTYERLVAQ FPSSGRFWKL YIEAEIKAKN YDKVEKLFQ RCLMKVLHID LWKCYLSYVR ETKGKLPSYK EKMAQAYDFA LDKIGMEIMS YQIWVDYINF LKGVEAVGSY A ENQRITAV ...String: MSGDGATEQA AEYVPEKVKK AEKKLEENPY DLDAWSILIR EAQNQPIDKA RKTYERLVAQ FPSSGRFWKL YIEAEIKAKN YDKVEKLFQ RCLMKVLHID LWKCYLSYVR ETKGKLPSYK EKMAQAYDFA LDKIGMEIMS YQIWVDYINF LKGVEAVGSY A ENQRITAV RRVYQRGCVN PMINIEQLWR DYNKYEEGIN IHLAKKMIED RSRDYMNARR VAKEYETVMK GLDRNAPSVP PQ NTPQEAQ QVDMWKKYIQ WEKSNPLRTE DQTLITKRVM FAYEQCLLVL GHHPDIWYEA AQYLEQSSKL LAEKGDMNNA KLF SDEAAN IYERAISTLL KKNMLLYFAY ADYEESRMKY EKVHSIYNRL LAIEDIDPTL VYIQYMKFAR RAEGIKSGRM IFKK AREDT RTRHHVYVTA ALMEYYCSKD KSVAFKIFEL GLKKYGDIPE YVLAYIDYLS HLNEDNNTRV LFERVLTSGS LPPEK SGEI WARFLAFESN IGDLASILKV EKRRFTAFKE EYEGKETALL VDRYKFMDLY PCSASELKAL GYKDVSRAKL AAIIPD PVV APSIVPVLKD EVDRKPEYPK PDTQQMIPFQ PRHLAPPGLH PVPGGVFPVP PAAVVLMKLL PPPICFQGPF VQVDELM EI FRRCKIPNTV EEAVRIITGG APELAVEGNG PVESNAVLTK AVKRPNEDSD EDEEKGAVVP PVHDIYRARQ QKRIR UniProtKB: Cleavage stimulation factor subunit 3 |
-Macromolecule #4: PAS RNA
Macromolecule | Name: PAS RNA / type: rna / ID: 4 / Number of copies: 1 |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 14.823655 KDa |
Sequence | String: UUCACAAAUA AACAUUUUUU UCACUGCAUU CUAGUUGUGG UUUGUCC |
-Macromolecule #6: ZINC ION
Macromolecule | Name: ZINC ION / type: ligand / ID: 6 / Number of copies: 3 / Formula: ZN |
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Molecular weight | Theoretical: 65.409 Da |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 0.25 mg/mL |
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Buffer | pH: 8 / Component - Concentration: 150.0 mM / Component - Formula: NaCl / Component - Name: sodium chloride / Details: 25 mM Tris-HCl, pH 8.0, 150 mM NaCl, 5 mM DTT |
Grid | Support film - Material: CARBON / Support film - topology: HOLEY / Details: unspecified |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV |
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Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Average exposure time: 10.0 sec. / Average electron dose: 70.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | C2 aperture diameter: 100.0 µm / Calibrated defocus max: 2.8000000000000003 µm / Calibrated defocus min: 0.9 µm / Calibrated magnification: 46729 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.2 µm / Nominal magnification: 225000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Image processing
Startup model | Type of model: INSILICO MODEL |
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Final reconstruction | Resolution.type: BY AUTHOR / Resolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION / Number images used: 50092 |
Initial angle assignment | Type: PROJECTION MATCHING |
Final angle assignment | Type: PROJECTION MATCHING |
-Atomic model buiding 1
Initial model |
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Output model | ![]() PDB-6uro: |
-Atomic model buiding 2
Initial model |
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Details | The model went through phenix.refine from the crystal structure(2OOE), but was not manually rebuilt due to lack of sufficient side chain density. | ||||||
Output model | ![]() PDB-6uro: |