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- EMDB-27265: CryoEM structure of human METTL1-WDR4 in complex with Lys-tRNA and SAM -

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Basic information

Entry
Database: EMDB / ID: EMD-27265
TitleCryoEM structure of human METTL1-WDR4 in complex with Lys-tRNA and SAM
Map dataRNA-protein complex
Sample
  • Complex: Human METTL1-WDR4 in complex with Lys-tRNA and SAM
    • Protein or peptide: tRNA (guanine-N(7)-)-methyltransferase non-catalytic subunit WDR4
    • Protein or peptide: tRNA (guanine-N(7)-)-methyltransferase
    • RNA: Lys-tRNA
  • Ligand: S-ADENOSYLMETHIONINES-Adenosyl methionine
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 / WD domain, G-beta repeat / WD40 repeats / WD40 repeat / Trp-Asp (WD) repeats profile. / Trp-Asp (WD) repeats circular profile. ...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 / 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
tRNA (guanine-N(7)-)-methyltransferase non-catalytic subunit WDR4 / tRNA (guanine-N(7)-)-methyltransferase
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.04 Å
AuthorsRuiz-Arroyo VM / Nam Y
Funding support United States, 3 items
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, 2022-
Header (metadata) releaseJan 11, 2023-
Map releaseJan 11, 2023-
UpdateJan 25, 2023-
Current statusJan 25, 2023Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_27265.png

Downloads & links

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Map

FileDownload / File: emd_27265.map.gz / Format: CCP4 / Size: 11.4 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationRNA-protein complex
Voxel sizeX=Y=Z: 1.26 Å
Density
Contour LevelBy AUTHOR: 0.168
Minimum - Maximum-0.14056808 - 0.4296963
Average (Standard dev.)0.0029826318 (±0.023364825)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions144144144
Spacing144144144
CellA=B=C: 181.44 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_27265_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Sharpen EM map.

Fileemd_27265_additional_1.map
AnnotationSharpen EM map.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: RNA-protein complex

Fileemd_27265_half_map_1.map
AnnotationRNA-protein complex
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: RNA-protein complex

Fileemd_27265_half_map_2.map
AnnotationRNA-protein complex
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 METTL1-WDR4 in complex with Lys-tRNA and SAM

EntireName: Human METTL1-WDR4 in complex with Lys-tRNA and SAM
Components
  • Complex: Human METTL1-WDR4 in complex with Lys-tRNA and SAM
    • Protein or peptide: tRNA (guanine-N(7)-)-methyltransferase non-catalytic subunit WDR4
    • Protein or peptide: tRNA (guanine-N(7)-)-methyltransferase
    • RNA: Lys-tRNA
  • Ligand: S-ADENOSYLMETHIONINES-Adenosyl methionine

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Supramolecule #1: Human METTL1-WDR4 in complex with Lys-tRNA and SAM

SupramoleculeName: Human METTL1-WDR4 in complex with Lys-tRNA and SAM / type: complex / ID: 1 / Chimera: Yes / Parent: 0 / Macromolecule list: #1-#3
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 112 KDa

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Macromolecule #1: tRNA (guanine-N(7)-)-methyltransferase non-catalytic subunit WDR4

MacromoleculeName: tRNA (guanine-N(7)-)-methyltransferase non-catalytic subunit WDR4
type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 45.123023 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MHHHHHHENL YFQGSGMAGS VGLALCGQTL VVRGGSRFLA TSIASSDDDS LFIYDCSAAE KKSQENKGED APLDQGSGAI LASTFSKSG SYFALTDDSK RLILFRTKPW QCLSVRTVAR RCTALTFIAS EEKVLVADKS GDVYSFSVLE PHGCGRLELG H LSMLLDVA ...String:
MHHHHHHENL YFQGSGMAGS VGLALCGQTL VVRGGSRFLA TSIASSDDDS LFIYDCSAAE KKSQENKGED APLDQGSGAI LASTFSKSG SYFALTDDSK RLILFRTKPW QCLSVRTVAR RCTALTFIAS EEKVLVADKS GDVYSFSVLE PHGCGRLELG H LSMLLDVA VSPDDRFILT ADRDEKIRVS WAAAPHSIES FCLGHTEFVS RISVVPTQPG LLLSSSGDGT LRLWEYRSGR QL HCCHLAS LQELVDPQAP QKFAASRIAF WCQENCVALL CDGTPVVYIF QLDARRQQLV YRQQLAFQHQ VWDVAFEETQ GLW VLQDCQ EAPLVLYRPV GDQWQSVPES TVLKKVSGVL RGNWAMLEGS AGADASFSSL YKATFDNVTS YLKKKEERLQ QQLE KKQRR

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Macromolecule #2: tRNA (guanine-N(7)-)-methyltransferase

MacromoleculeName: tRNA (guanine-N(7)-)-methyltransferase / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO / EC number: tRNA (guanine46-N7)-methyltransferase
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 31.472004 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MAAETRNVAG AEAPPPQKRY YRQRAHSNPM ADHTLRYPVK PEEMDWSELY PEFFAPLTQN QSHDDPKDKK EKRAQAQVEF ADIGCGYGG LLVELSPLFP DTLILGLEIR VKVSDYVQDR IRALRAAPAG GFQNIACLRS NAMKHLPNFF YKGQLTKMFF L FPAPHFKR ...String:
MAAETRNVAG AEAPPPQKRY YRQRAHSNPM ADHTLRYPVK PEEMDWSELY PEFFAPLTQN QSHDDPKDKK EKRAQAQVEF ADIGCGYGG LLVELSPLFP DTLILGLEIR VKVSDYVQDR IRALRAAPAG GFQNIACLRS NAMKHLPNFF YKGQLTKMFF L FPAPHFKR TKHKWRIISP TLLAEYAYVL RVGGLVYTIT DVLELHDWMC THFEEHPLFE RVPLEDLSED PVVGHLGTST EE GKKVLRN GGKNFPAIFR RIQDPVLQAV TSQTSLPGH

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Macromolecule #3: Lys-tRNA

MacromoleculeName: Lys-tRNA / type: rna / ID: 3 / Number of copies: 1
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 23.507912 KDa
SequenceString:
GCCCGGAUAG CUCAGUCGGU AGAGCAUCAG ACUUUUAAUC UGAGGGUCCA GGGUUCAAGU CCCUGUUCGG GCG

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Macromolecule #4: S-ADENOSYLMETHIONINE

MacromoleculeName: S-ADENOSYLMETHIONINE / type: ligand / ID: 4 / Number of copies: 1 / Formula: SAM
Molecular weightTheoretical: 398.437 Da
Chemical component information

ChemComp-SAM:
S-ADENOSYLMETHIONINE / S-Adenosyl methionine

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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

Concentration3 mg/mL
BufferpH: 7.5
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 273.15 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.2 µm / Nominal defocus min: 0.7000000000000001 µm
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 62.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.3.1)
Final 3D classificationSoftware - Name: cryoSPARC (ver. 3.3.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.3.1)
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 4.04 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.3.1) / Number images used: 34558
FSC plot (resolution estimation)

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