|Entry||Database: PDB / ID: 5zya|
|Title||SF3b spliceosomal complex bound to E7107|
|Keywords||SPLICING / Splicing Modulator / SF3b sub-complex|
|Function / homology||Mono-functional DNA-alkylating methyl methanesulfonate N-term / Splicing factor 3B, subunit 5 / PHF5-like / Splicing factor 3B subunit 5/RDS3 complex subunit 10 / Armadillo-like helical / Splicing factor 3B subunit 1 / WD40/YVTN repeat-like-containing domain superfamily / Armadillo-type fold / WD40-repeat-containing domain superfamily / Splicing factor 3B subunit 1-like ...Mono-functional DNA-alkylating methyl methanesulfonate N-term / Splicing factor 3B, subunit 5 / PHF5-like / Splicing factor 3B subunit 5/RDS3 complex subunit 10 / Armadillo-like helical / Splicing factor 3B subunit 1 / WD40/YVTN repeat-like-containing domain superfamily / Armadillo-type fold / WD40-repeat-containing domain superfamily / Splicing factor 3B subunit 1-like / CPSF A subunit region / PHF5-like protein / Splicing factor 3B subunit 10 (SF3b10) / Splicing factor 3B subunit 1 / B-WICH complex positively regulates rRNA expression / mRNA Splicing - Major Pathway / mRNA Splicing - Minor Pathway / Cleavage/polyadenylation specificity factor, A subunit, C-terminal / U11/U12 snRNP / RNA splicing, via transesterification reactions / U2-type precatalytic spliceosome / small nuclear ribonucleoprotein complex / U12-type spliceosomal complex / U2 snRNP / positive regulation of gene expression, epigenetic / spliceosomal complex assembly / U2-type prespliceosome / catalytic step 2 spliceosome / spliceosomal complex / mRNA splicing, via spliceosome / negative regulation of protein catabolic process / nuclear matrix / nucleic acid binding / mRNA binding / protein-containing complex binding / nuclear speck / nucleolus / DNA-binding transcription factor activity / positive regulation of transcription, DNA-templated / RNA binding / DNA binding / nucleoplasm / nucleus / metal ion binding / Splicing factor 3B subunit 1 / Splicing factor 3B subunit 3 / PHD finger-like domain-containing protein 5A / Splicing factor 3B subunit 5|
Function and homology information
|Specimen source||Homo sapiens (human)|
|Method||ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / 3.95 Å resolution|
|Authors||Finci, L.I. / Larsen, N.A.|
|Citation||Journal: Genes Dev. / Year: 2018|
Title: The cryo-EM structure of the SF3b spliceosome complex bound to a splicing modulator reveals a pre-mRNA substrate competitive mechanism of action.
Authors: Lorenzo I Finci / Xiaofeng Zhang / Xiuliang Huang / Qiang Zhou / Jennifer Tsai / Teng Teng / Anant Agrawal / Betty Chan / Sean Irwin / Craig Karr / Andrew Cook / Ping Zhu / Dominic Reynolds / Peter G Smith / Peter Fekkes / Silvia Buonamici / Nicholas A Larsen
Abstract: Somatic mutations in spliceosome proteins lead to dysregulated RNA splicing and are observed in a variety of cancers. These genetic aberrations may offer a potential intervention point for targeted ...Somatic mutations in spliceosome proteins lead to dysregulated RNA splicing and are observed in a variety of cancers. These genetic aberrations may offer a potential intervention point for targeted therapeutics. SF3B1, part of the U2 small nuclear RNP (snRNP), is targeted by splicing modulators, including E7107, the first to enter clinical trials, and, more recently, H3B-8800. Modulating splicing represents a first-in-class opportunity in drug discovery, and elucidating the structural basis for the mode of action opens up new possibilities for structure-based drug design. Here, we present the cryogenic electron microscopy (cryo-EM) structure of the SF3b subcomplex (SF3B1, SF3B3, PHF5A, and SF3B5) bound to E7107 at 3.95 Å. This structure shows that E7107 binds in the branch point adenosine-binding pocket, forming close contacts with key residues that confer resistance upon mutation: SF3B1 and PHF5A The structure suggests a model in which splicing modulators interfere with branch point adenosine recognition and supports a substrate competitive mechanism of action (MOA). Using several related chemical probes, we validate the pose of the compound and support their substrate competitive MOA by comparing their activity against both strong and weak pre-mRNA substrates. Finally, we present functional data and structure-activity relationship (SAR) on the PHF5A mutation that sensitizes cells to some chemical probes but not others. Developing small molecule splicing modulators represents a promising therapeutic approach for a variety of diseases, and this work provides a significant step in enabling structure-based drug design for these elaborate natural products. Importantly, this work also demonstrates that the utilization of cryo-EM in drug discovery is coming of age.
SummaryFull reportAbout validation report
|Date||Deposition: May 23, 2018 / Release: Jun 20, 2018|
|Structure viewer||Molecule: |
Downloads & links
B: Splicing factor 3B subunit 5
C: Splicing factor 3B subunit 1
D: PHD finger-like domain-containing protein 5A
A: Splicing factor 3B subunit 3
-Splicing factor 3B subunit ... , 3 types, 3 molecules B
|#1: Protein/peptide|| |
Mass: 10149.369 Da / Num. of mol.: 1 / Source: (gene. exp.) Homo sapiens (human) / Gene: SF3B5 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q9BWJ5
|#2: Protein/peptide|| |
Mass: 146024.938 Da / Num. of mol.: 1 / Source: (gene. exp.) Homo sapiens (human) / Gene: SF3B1 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: O75533
|#4: Protein/peptide|| |
Mass: 136471.562 Da / Num. of mol.: 1 / Source: (gene. exp.) Homo sapiens (human) / Gene: SF3B3 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q15393
-Protein/peptide , 1 types, 1 molecules D
|#3: Protein/peptide|| |
Mass: 9394.955 Da / Num. of mol.: 1 / Source: (gene. exp.) Homo sapiens (human) / Gene: PHF5A / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q7RTV0
-Non-polymers , 3 types, 5 molecules
|#5: Chemical|| ChemComp-9B0 / [(|
|#6: Chemical||#7: Chemical|| ChemComp-K / |
|Experiment||Method: ELECTRON MICROSCOPY|
|EM experiment||Aggregation state: PARTICLE / Reconstruction method: single particle reconstruction|
|Component||Name: SF3b Sub-complex / Type: COMPLEX / Entity ID: 1,||Source (natural)||Organism: Homo sapiens (human)||Source (recombinant)||Organism: Spodoptera frugiperda (fall armyworm)||Buffer solution||pH: 8||Specimen||Conc.: 1 / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES||Vitrification||Cryogen name: ETHANE|
-Electron microscopy imaging
Model: Titan Krios / Image courtesy: FEI Company
|Microscopy||Microscope model: FEI TITAN KRIOS|
|Electron gun||Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 / Illumination mode: FLOOD BEAM|
|Electron lens||Mode: BRIGHT FIELDBright-field microscopy|
|Image recording||Electron dose: 55 / Film or detector model: GATAN K2 SUMMIT (4k x 4k)|
|Software||Name: PHENIX / Version: 1.11.1_2575: / Classification: refinement|
|CTF correction||Type: NONE|
|3D reconstruction||Resolution: 3.95 / Resolution method: FSC 0.143 CUT-OFF / Number of particles: 241288 / Symmetry type: POINT|
|Least-squares process||Highest resolution: 3.95|
|Refine LS restraints|
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