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- PDB-8d9l: CryoEM structure of human METTL1-WDR4 in complex with Lys-tRNA and SAM -

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Basic information

Entry
Database: PDB / ID: 8d9l
TitleCryoEM structure of human METTL1-WDR4 in complex with Lys-tRNA and SAM
Components
  • Lys-tRNA
  • tRNA (guanine-N(7)-)-methyltransferase
  • tRNA (guanine-N(7)-)-methyltransferase non-catalytic subunit WDR4
KeywordsTRANSFERASE/RNA / Cancer protein / Methyl transferase / TRANSFERASE / TRANSFERASE-RNA complex
Function / homology
Function and homology information


internal mRNA (guanine-N7-)-methyltransferase activity / tRNA stabilization / tRNA (m7G46) methyltransferase complex / tRNA (guanine-N7)-methylation / RNA (guanine-N7)-methylation / tRNA (guanine46-N7)-methyltransferase / tRNA (guanine(46)-N7)-methyltransferase activity / tRNA methyltransferase activator activity / tRNA methyltransferase complex / tRNA modification in the nucleus and cytosol ...internal mRNA (guanine-N7-)-methyltransferase activity / tRNA stabilization / tRNA (m7G46) methyltransferase complex / tRNA (guanine-N7)-methylation / RNA (guanine-N7)-methylation / tRNA (guanine46-N7)-methyltransferase / tRNA (guanine(46)-N7)-methyltransferase activity / tRNA methyltransferase activator activity / tRNA methyltransferase complex / tRNA modification in the nucleus and cytosol / tRNA methylation / tRNA modification / enzyme activator activity / Transferases; Transferring one-carbon groups; Methyltransferases / chromosome / tRNA binding / DNA damage response / nucleolus / nucleoplasm / nucleus / cytosol
Similarity search - Function
tRNA (guanine-N(7)-)-methyltransferase non-catalytic subunit / tRNA (guanine-N-7) methyltransferase catalytic subunit Trm8, eukaryote / SAM-dependent methyltransferase TRMB-type domain profile. / tRNA (guanine-N-7) methyltransferase, Trmb type / Putative methyltransferase / Trp-Asp (WD) repeats profile. / Trp-Asp (WD) repeats circular profile. / WD domain, G-beta repeat / WD40 repeats / WD40 repeat ...tRNA (guanine-N(7)-)-methyltransferase non-catalytic subunit / tRNA (guanine-N-7) methyltransferase catalytic subunit Trm8, eukaryote / SAM-dependent methyltransferase TRMB-type domain profile. / tRNA (guanine-N-7) methyltransferase, Trmb type / Putative methyltransferase / Trp-Asp (WD) repeats profile. / Trp-Asp (WD) repeats circular profile. / WD domain, G-beta repeat / WD40 repeats / WD40 repeat / WD40-repeat-containing domain superfamily / WD40/YVTN repeat-like-containing domain superfamily / S-adenosyl-L-methionine-dependent methyltransferase superfamily
Similarity search - Domain/homology
S-ADENOSYLMETHIONINE / : / RNA / RNA (> 10) / tRNA (guanine-N(7)-)-methyltransferase non-catalytic subunit WDR4 / tRNA (guanine-N(7)-)-methyltransferase
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.04 Å
AuthorsRuiz-Arroyo, V.M. / Nam, Y.
Funding support United States, 3items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01 GM122960 United States
Cancer Prevention and Research Institute of Texas (CPRIT)RP190259 United States
Welch FoundationI-2115-20220331 United States
CitationJournal: Nature / Year: 2023
Title: Structures and mechanisms of tRNA methylation by METTL1-WDR4.
Authors: Victor M Ruiz-Arroyo / Rishi Raj / Kesavan Babu / Otgonbileg Onolbaatar / Paul H Roberts / Yunsun Nam /
Abstract: Specific, regulated modification of RNAs is important for proper gene expression. tRNAs are rich with various chemical modifications that affect their stability and function. 7-Methylguanosine (mG) ...Specific, regulated modification of RNAs is important for proper gene expression. tRNAs are rich with various chemical modifications that affect their stability and function. 7-Methylguanosine (mG) at tRNA position 46 is a conserved modification that modulates steady-state tRNA levels to affect cell growth. The METTL1-WDR4 complex generates mG46 in humans, and dysregulation of METTL1-WDR4 has been linked to brain malformation and multiple cancers. Here we show how METTL1 and WDR4 cooperate to recognize RNA substrates and catalyse methylation. A crystal structure of METTL1-WDR4 and cryo-electron microscopy structures of METTL1-WDR4-tRNA show that the composite protein surface recognizes the tRNA elbow through shape complementarity. The cryo-electron microscopy structures of METTL1-WDR4-tRNA with S-adenosylmethionine or S-adenosylhomocysteine along with METTL1 crystal structures provide additional insights into the catalytic mechanism by revealing the active site in multiple states. The METTL1 N terminus couples cofactor binding with conformational changes in the tRNA, the catalytic loop and the WDR4 C terminus, acting as the switch to activate mG methylation. Thus, our structural models explain how post-translational modifications of the METTL1 N terminus can regulate methylation. Together, our work elucidates the core and regulatory mechanisms underlying mG modification by METTL1, providing the framework to understand its contribution to biology and disease.
History
DepositionJun 10, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jan 11, 2023Provider: repository / Type: Initial release
Revision 1.1Jan 25, 2023Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation_author.identifier_ORCID
Revision 1.2Jun 12, 2024Group: Data collection / Category: chem_comp_atom / chem_comp_bond

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
B: tRNA (guanine-N(7)-)-methyltransferase non-catalytic subunit WDR4
A: tRNA (guanine-N(7)-)-methyltransferase
C: Lys-tRNA
hetero molecules


Theoretical massNumber of molelcules
Total (without water)100,5014
Polymers100,1033
Non-polymers3981
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein tRNA (guanine-N(7)-)-methyltransferase non-catalytic subunit WDR4 / Protein Wuho homolog / hWH / WD repeat-containing protein 4


Mass: 45123.023 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: WDR4 / Production host: Escherichia coli (E. coli) / References: UniProt: P57081
#2: Protein tRNA (guanine-N(7)-)-methyltransferase / Methyltransferase-like protein 1 / mRNA (guanine-N(7)-)-methyltransferase / miRNA (guanine-N(7)-)- ...Methyltransferase-like protein 1 / mRNA (guanine-N(7)-)-methyltransferase / miRNA (guanine-N(7)-)-methyltransferase / tRNA (guanine(46)-N(7))-methyltransferase / tRNA(m7G46)-methyltransferase


Mass: 31472.004 Da / Num. of mol.: 1 / Mutation: D163A
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: METTL1, C12orf1 / Production host: Escherichia coli (E. coli)
References: UniProt: Q9UBP6, tRNA (guanine46-N7)-methyltransferase, Transferases; Transferring one-carbon groups; Methyltransferases
#3: RNA chain Lys-tRNA


Mass: 23507.912 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Homo sapiens (human) / References: GenBank: 1781322811
#4: Chemical ChemComp-SAM / S-ADENOSYLMETHIONINE


Mass: 398.437 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C15H22N6O5S / 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: Human METTL1-WDR4 in complex with Lys-tRNA and SAM / Type: COMPLEX / Entity ID: #1-#3 / Source: RECOMBINANT
Molecular weightValue: 0.112 MDa / Experimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 7.5
SpecimenConc.: 3 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid mesh size: 200 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 273.15 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 FIELD / Nominal defocus max: 2200 nm / Nominal defocus min: 700 nm
Image recordingElectron dose: 62 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

Software
NameVersionClassification
phenix.real_space_refine1.19.2_4158refinement
PHENIX1.19.2_4158refinement
EM software
IDNameVersionCategory
4cryoSPARC3.3.1CTF correction
9cryoSPARC3.3.1initial Euler assignment
10cryoSPARC3.3.1final Euler assignment
11cryoSPARC3.3.1classification
12cryoSPARC3.3.13D reconstruction
CTF correctionType: NONE
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 4.04 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 34558 / Symmetry type: POINT
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 267.66 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00136026
ELECTRON MICROSCOPYf_angle_d0.38358487
ELECTRON MICROSCOPYf_chiral_restr0.03441001
ELECTRON MICROSCOPYf_plane_restr0.0023837
ELECTRON MICROSCOPYf_dihedral_angle_d9.7192382

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