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- PDB-7m05: CryoEM structure of PRMT5 bound to covalent PBM-site inhibitor BR... -

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Basic information

Entry
Database: PDB / ID: 7m05
TitleCryoEM structure of PRMT5 bound to covalent PBM-site inhibitor BRD-6988
Components
  • Methylosome protein 50WD repeat-containing protein 77
  • Protein arginine N-methyltransferase 5
KeywordsTRANSFERASE/INHIBITOR / methyltransferase / splicing / epigenetic / TRANSFERASE-INHIBITOR complex
Function / homology
Function and homology information


positive regulation of adenylate cyclase-inhibiting dopamine receptor signaling pathway / peptidyl-arginine N-methylation / oocyte axis specification / type II protein arginine methyltransferase / protein-arginine omega-N symmetric methyltransferase activity / peptidyl-arginine methylation / Golgi ribbon formation / negative regulation of epithelial cell proliferation involved in prostate gland development / histone H4R3 methyltransferase activity / secretory columnal luminar epithelial cell differentiation involved in prostate glandular acinus development ...positive regulation of adenylate cyclase-inhibiting dopamine receptor signaling pathway / peptidyl-arginine N-methylation / oocyte axis specification / type II protein arginine methyltransferase / protein-arginine omega-N symmetric methyltransferase activity / peptidyl-arginine methylation / Golgi ribbon formation / negative regulation of epithelial cell proliferation involved in prostate gland development / histone H4R3 methyltransferase activity / secretory columnal luminar epithelial cell differentiation involved in prostate glandular acinus development / epithelial cell proliferation involved in prostate gland development / histone arginine N-methyltransferase activity / methylosome / protein-arginine N-methyltransferase activity / positive regulation of mRNA splicing, via spliceosome / methyl-CpG binding / : / endothelial cell activation / histone H3 methyltransferase activity / Cul4B-RING E3 ubiquitin ligase complex / histone methyltransferase complex / regulation of mitotic nuclear division / positive regulation of oligodendrocyte differentiation / histone methyltransferase activity / E-box binding / negative regulation of cell differentiation / ubiquitin-like ligase-substrate adaptor activity / spliceosomal snRNP assembly / ribonucleoprotein complex binding / regulation of ERK1 and ERK2 cascade / nuclear receptor coactivator activity / regulation of signal transduction by p53 class mediator / liver regeneration / methyltransferase activity / DNA-templated transcription termination / circadian regulation of gene expression / Regulation of TP53 Activity through Methylation / RMTs methylate histone arginines / protein polyubiquitination / transcription corepressor activity / p53 binding / snRNP Assembly / ubiquitin-dependent protein catabolic process / chromatin remodeling / protein heterodimerization activity / positive regulation of cell population proliferation / chromatin / regulation of DNA-templated transcription / regulation of transcription by RNA polymerase II / Golgi apparatus / nucleoplasm / identical protein binding / nucleus / cytosol / cytoplasm
Similarity search - Function
Protein arginine N-methyltransferase PRMT5 / PRMT5 arginine-N-methyltransferase / PRMT5, TIM barrel domain / PRMT5, oligomerisation domain / PRMT5 arginine-N-methyltransferase / PRMT5 TIM barrel domain / PRMT5 oligomerisation domain / Protein arginine N-methyltransferase / SAM-dependent methyltransferase PRMT-type domain profile. / WD40 repeat, conserved site ...Protein arginine N-methyltransferase PRMT5 / PRMT5 arginine-N-methyltransferase / PRMT5, TIM barrel domain / PRMT5, oligomerisation domain / PRMT5 arginine-N-methyltransferase / PRMT5 TIM barrel domain / PRMT5 oligomerisation domain / Protein arginine N-methyltransferase / SAM-dependent methyltransferase PRMT-type domain profile. / WD40 repeat, conserved site / Trp-Asp (WD) repeats signature. / 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 / S-adenosyl-L-methionine-dependent methyltransferase superfamily
Similarity search - Domain/homology
Chem-YJG / Protein arginine N-methyltransferase 5 / Methylosome protein WDR77
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.39 Å
AuthorsMcMillan, B.J. / McKinney, D.C. / Timm, D.E.
Citation
Journal: J Med Chem / Year: 2021
Title: Discovery of a First-in-Class Inhibitor of the PRMT5-Substrate Adaptor Interaction.
Authors: David C McKinney / Brian J McMillan / Matthew J Ranaghan / Jamie A Moroco / Merissa Brousseau / Zachary Mullin-Bernstein / Meghan O'Keefe / Patrick McCarren / Michael F Mesleh / Kathleen M ...Authors: David C McKinney / Brian J McMillan / Matthew J Ranaghan / Jamie A Moroco / Merissa Brousseau / Zachary Mullin-Bernstein / Meghan O'Keefe / Patrick McCarren / Michael F Mesleh / Kathleen M Mulvaney / Foxy Robinson / Ritu Singh / Besnik Bajrami / Florence F Wagner / Robert Hilgraf / Martin J Drysdale / Arthur J Campbell / Adam Skepner / David E Timm / Dale Porter / Virendar K Kaushik / William R Sellers / Alessandra Ianari /
Abstract: PRMT5 and its substrate adaptor proteins (SAPs), pICln and Riok1, are synthetic lethal dependencies in MTAP-deleted cancer cells. SAPs share a conserved PRMT5 binding motif (PBM) which mediates ...PRMT5 and its substrate adaptor proteins (SAPs), pICln and Riok1, are synthetic lethal dependencies in MTAP-deleted cancer cells. SAPs share a conserved PRMT5 binding motif (PBM) which mediates binding to a surface of PRMT5 distal to the catalytic site. This interaction is required for methylation of several PRMT5 substrates, including histone and spliceosome complexes. We screened for small molecule inhibitors of the PRMT5-PBM interaction and validated a compound series which binds to the PRMT5-PBM interface and directly inhibits binding of SAPs. Mode of action studies revealed the formation of a covalent bond between a halogenated pyridazinone group and cysteine 278 of PRMT5. Optimization of the starting hit produced a lead compound, BRD0639, which engages the target in cells, disrupts PRMT5-RIOK1 complexes, and reduces substrate methylation. BRD0639 is a first-in-class PBM-competitive inhibitor that can support studies of PBM-dependent PRMT5 activities and the development of novel PRMT5 inhibitors that selectively target these functions.
#1: Journal: bioRxiv / Year: 2020
Title: Discovery of a first-in-class inhibitor of the PRMT5-substrate adaptor interaction
Authors: Mulvaney, K.M. / McMillan, B.J. / Sellers, W.R.
History
DepositionMar 10, 2021Deposition site: RCSB / Processing site: RCSB
Revision 1.0Mar 17, 2021Provider: repository / Type: Initial release
Revision 1.1Aug 25, 2021Group: Database references / Category: citation / citation_author / database_2
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession

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Structure visualization

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Assembly

Deposited unit
A: Protein arginine N-methyltransferase 5
B: Methylosome protein 50
C: Protein arginine N-methyltransferase 5
D: Methylosome protein 50
E: Protein arginine N-methyltransferase 5
F: Methylosome protein 50
G: Protein arginine N-methyltransferase 5
H: Methylosome protein 50
hetero molecules


Theoretical massNumber of molelcules
Total (without water)439,80712
Polymers437,9598
Non-polymers1,8484
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: equilibrium centrifugation
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein
Protein arginine N-methyltransferase 5 / 72 kDa ICln-binding protein / Histone-arginine N-methyltransferase PRMT5 / Jak-binding protein 1 / ...72 kDa ICln-binding protein / Histone-arginine N-methyltransferase PRMT5 / Jak-binding protein 1 / Shk1 kinase-binding protein 1 homolog / SKB1Hs


Mass: 72766.664 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: PRMT5, HRMT1L5, IBP72, JBP1, SKB1 / Production host: Spodoptera frugiperda (fall armyworm)
References: UniProt: O14744, type II protein arginine methyltransferase
#2: Protein
Methylosome protein 50 / WD repeat-containing protein 77 / MEP-50 / Androgen receptor cofactor p44 / WD repeat-containing protein 77 / p44/Mep50


Mass: 36723.164 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: WDR77, MEP50, WD45, HKMT1069, Nbla10071 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q9BQA1
#3: Chemical
ChemComp-YJG / 2-(5-chloro-6-oxopyridazin-1(6H)-yl)-N-(4-methyl-3-{[2-(pyridin-2-yl)ethyl]sulfamoyl}phenyl)acetamide


Mass: 461.922 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C20H20ClN5O4S / Feature type: SUBJECT OF INVESTIGATION
Has ligand of interestY

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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: Hetero-octamer complex of PRMT5 and WDR77 / Type: COMPLEX / Entity ID: #1-#2 / Source: RECOMBINANT
Molecular weightValue: 0.437 MDa / Experimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Spodoptera frugiperda (fall armyworm)
Buffer solutionpH: 7.4
Buffer component
IDConc.NameFormulaBuffer-ID
110 mMHEPES1
2150 mMsodium chlorideNaClSodium chloride1
31 mMTCEP1
450 nMJNJ-646191781
SpecimenConc.: 1.42 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: UltrAuFoil
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277.15 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 130000 X / Calibrated magnification: 46296 X / Nominal defocus max: 2100 nm / Nominal defocus min: 1400 nm / Cs: 2.7 mm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 8 sec. / Electron dose: 62.4 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 2169
EM imaging opticsEnergyfilter name: GIF Quantum LS / Energyfilter slit width: 30 eV
Image scansSampling size: 5 µm / Width: 3838 / Height: 3710 / Movie frames/image: 40 / Used frames/image: 4-40

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Processing

SoftwareName: PHENIX / Version: 1.16_3549: / Classification: refinement
EM software
IDNameVersionCategory
1cisTEM1particle selection
2EPU2.4.0.47RELimage acquisition
4cisTEM1CTF correction
7PHENIX1.16-3549model fitting
9PHENIX1.16-3549model refinement
10cisTEM1initial Euler assignment
11cisTEM1final Euler assignment
12cisTEM1classification
13cisTEM13D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 956646
SymmetryPoint symmetry: D2 (2x2 fold dihedral)
3D reconstructionResolution: 2.39 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 443624 / Algorithm: FOURIER SPACE / Num. of class averages: 22 / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT / Space: REAL
Atomic model buildingPDB-ID: 6V0P

6v0p

Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0158040
ELECTRON MICROSCOPYf_angle_d0.843104804
ELECTRON MICROSCOPYf_dihedral_angle_d10.82923452
ELECTRON MICROSCOPYf_chiral_restr0.0584448
ELECTRON MICROSCOPYf_plane_restr0.0058652

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