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- EMDB-33854: Cryo-EM structure of SAM-bound MTA1-MTA9-p1-p2 complex -

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Basic information

Entry
Database: EMDB / ID: EMD-33854
TitleCryo-EM structure of SAM-bound MTA1-MTA9-p1-p2 complex
Map data
Sample
  • Complex: MTAc holoenzyme
    • Protein or peptide: MTA9
    • Protein or peptide: MT-a70 family protein
    • Protein or peptide: P1
    • Protein or peptide: Transmembrane protein, putative
  • Ligand: S-ADENOSYLMETHIONINE
KeywordsN6-adenine methylation / MTAc holoenzyme / DNA BINDING PROTEIN
Function / homology
Function and homology information


RNA N6-methyladenosine methyltransferase complex / : / methyltransferase activity / membrane / nucleus
Similarity search - Function
MT-A70-like / MT-A70 / MT-A70-like family profile. / Myb-like DNA-binding domain / SANT/Myb domain / Homeobox-like domain superfamily / S-adenosyl-L-methionine-dependent methyltransferase superfamily
Similarity search - Domain/homology
Transmembrane protein, putative / Uncharacterized protein / MT-a70 family protein / Myb-like domain-containing protein
Similarity search - Component
Biological speciesTetrahymena thermophila SB210 (eukaryote)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.6 Å
AuthorsYan JJ / Guan ZY / Liu FQ / Yan XH / Hou MJ / Yin P
Funding support China, 1 items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC) China
CitationJournal: Cell Discov / Year: 2023
Title: Structural insights into DNA N-adenine methylation by the MTA1 complex.
Authors: Junjun Yan / Feiqing Liu / Zeyuan Guan / Xuhui Yan / Xiaohuan Jin / Qiang Wang / Zican Wang / Junjie Yan / Delin Zhang / Zhu Liu / Shan Wu / Ping Yin /
Abstract: N-methyldeoxyadenine (6mA) has recently been reported as a prevalent DNA modification in eukaryotes. The Tetrahymena thermophila MTA1 complex consisting of four subunits, namely MTA1, MTA9, p1, and ...N-methyldeoxyadenine (6mA) has recently been reported as a prevalent DNA modification in eukaryotes. The Tetrahymena thermophila MTA1 complex consisting of four subunits, namely MTA1, MTA9, p1, and p2, is the first identified eukaryotic 6mA methyltransferase (MTase) complex. Unlike the prokaryotic 6mA MTases which have been biochemically and structurally characterized, the operation mode of the MTA1 complex remains largely elusive. Here, we report the cryogenic electron microscopy structures of the quaternary MTA1 complex in S-adenosyl methionine (SAM)-bound (2.6 Å) and S-adenosyl homocysteine (SAH)-bound (2.8 Å) states. Using an AI-empowered integrative approach based on AlphaFold prediction and chemical cross-linking mass spectrometry, we further modeled a near-complete structure of the quaternary complex. Coupled with biochemical characterization, we revealed that MTA1 serves as the catalytic core, MTA1, MTA9, and p1 likely accommodate the substrate DNA, and p2 may facilitate the stabilization of MTA1. These results together offer insights into the molecular mechanism underpinning methylation by the MTA1 complex and the potential diversification of MTases for N-adenine methylation.
History
DepositionJul 15, 2022-
Header (metadata) releaseJan 18, 2023-
Map releaseJan 18, 2023-
UpdateAug 2, 2023-
Current statusAug 2, 2023Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_33854.png

Downloads & links

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Map

FileDownload / File: emd_33854.map.gz / Format: CCP4 / Size: 52.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 0.85 Å
Density
Contour LevelBy AUTHOR: 0.35
Minimum - Maximum-2.3308094 - 4.036278
Average (Standard dev.)-0.00035666735 (±0.07634106)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions240240240
Spacing240240240
CellA=B=C: 204.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_33854_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_33854_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : MTAc holoenzyme

EntireName: MTAc holoenzyme
Components
  • Complex: MTAc holoenzyme
    • Protein or peptide: MTA9
    • Protein or peptide: MT-a70 family protein
    • Protein or peptide: P1
    • Protein or peptide: Transmembrane protein, putative
  • Ligand: S-ADENOSYLMETHIONINE

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Supramolecule #1: MTAc holoenzyme

SupramoleculeName: MTAc holoenzyme / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#4
Source (natural)Organism: Tetrahymena thermophila SB210 (eukaryote)

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Macromolecule #1: MTA9

MacromoleculeName: MTA9 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Tetrahymena thermophila SB210 (eukaryote) / Strain: SB210
Molecular weightTheoretical: 52.026379 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MAPKKQEQEP IRLSTRTASK KVDYLQLSNG KLEDFFDDLE EDNKPARNRS RSKKRGRKPL KKADSRSKTP SRVSNARGRS KSLGPRKTY PRKKNLSPDN QLSLLLKWRN DKIPLKSASE TDNKCKVVNV KNIFKSDLSK YGANLQALFI NALWKVKSRK E KEGLNIND ...String:
MAPKKQEQEP IRLSTRTASK KVDYLQLSNG KLEDFFDDLE EDNKPARNRS RSKKRGRKPL KKADSRSKTP SRVSNARGRS KSLGPRKTY PRKKNLSPDN QLSLLLKWRN DKIPLKSASE TDNKCKVVNV KNIFKSDLSK YGANLQALFI NALWKVKSRK E KEGLNIND LSNLKIPLSL MKNGILFIWS EKEILGQIVE IMEQKGFTYI ENFSIMFLGL NKCLQSINHK DEDSQNSTAS TN NTNNEAI TSDLTLKDTS KFSDQIQDNH SEDSDQARKQ QTPDDITQKK NKLLKKSSVP SIQKLFEEDP VQTPSVNKPI EKS IEQVTQ EKKFVMNNLD ILKSTDINNL FLRNNYPYFK KTRHTLLMFR RIGDKNQKLE LRHQRTSDVV FEVTDEQDPS KVDT MMKEY VYQMIETLLP KAQFIPGVDK HLKMMELFAS TDNYRPGWIS VIEK

UniProtKB: Uncharacterized protein

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Macromolecule #2: MT-a70 family protein

MacromoleculeName: MT-a70 family protein / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Tetrahymena thermophila SB210 (eukaryote) / Strain: SB210
Molecular weightTheoretical: 42.696059 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MSKAVNKKGL RPRKSDSILD HIKNKLDQEF LEDNENGEQS DEDYDQKSLN KAKKPYKKRQ TQNGSELVIS QQKTKAKASA NNKKSAKNS QKLDEEEKIV EEEDLSPQKN GAVSEDDQQQ EASTQEDDYL DRLPKSKKGL QGLLQDIEKR ILHYKQLFFK E QNEIANGK ...String:
MSKAVNKKGL RPRKSDSILD HIKNKLDQEF LEDNENGEQS DEDYDQKSLN KAKKPYKKRQ TQNGSELVIS QQKTKAKASA NNKKSAKNS QKLDEEEKIV EEEDLSPQKN GAVSEDDQQQ EASTQEDDYL DRLPKSKKGL QGLLQDIEKR ILHYKQLFFK E QNEIANGK RSMVPDNSIP ICSDVTKLNF QALIDAQMRH AGKMFDVIMM DPPWQLSSSQ PSRGVAIAYD SLSDEKIQNM PI QSLQQDG FIFVWAINAK YRVTIKMIEN WGYKLVDEIT WVKKTVNGKI AKGHGFYLQH AKESCLIGVK GDVDNGRFKK NIA SDVIFS ERRGQSQKPE EIYQYINQLC PNGNYLEIFA RRNNLHDNWV SIGNEL

UniProtKB: MT-a70 family protein

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Macromolecule #3: P1

MacromoleculeName: P1 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Tetrahymena thermophila SB210 (eukaryote) / Strain: SB210
Molecular weightTheoretical: 41.602758 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MSLKKGKFQH NQSKSLWNYT LSPGWREEEV KILKSALQLF GIGKWKKIME SGCLPGKSIG QIYMQTQRLL GQQSLGDFMG LQIDLEAVF NQNMKKQDVL RKNNCIINTG DNPTKEERKR RIEQNRKIYG LSAKQIAEIK LPKVKKHAPQ YMTLEDIENE K FTNLEILT ...String:
MSLKKGKFQH NQSKSLWNYT LSPGWREEEV KILKSALQLF GIGKWKKIME SGCLPGKSIG QIYMQTQRLL GQQSLGDFMG LQIDLEAVF NQNMKKQDVL RKNNCIINTG DNPTKEERKR RIEQNRKIYG LSAKQIAEIK LPKVKKHAPQ YMTLEDIENE K FTNLEILT HLYNLKAEIV RRLAEQGETI AQPSIIKSLN NLNHNLEQNQ NSNSSTETKV TLEQSGKKKY KVLAIEETEL QN GPIATNS QKKSINGKRK NNRKINSDSE GNEEDISLED IDSQESEINS EEIVEDDEED EQIEEPSKIK KRKKNPEQES EED DIEEDQ EEDELVVNEE EIFEDDDDDE DNQDSSEDDD DDED

UniProtKB: Myb-like domain-containing protein

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Macromolecule #4: Transmembrane protein, putative

MacromoleculeName: Transmembrane protein, putative / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Tetrahymena thermophila SB210 (eukaryote) / Strain: SB210
Molecular weightTheoretical: 19.886547 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString:
MKHHHHHHHG AAGTSLYKKA GENLYFQGSM KKNGKSQNQP LDFTQYAKNM RKDLSNQDIC LEDGALNHSY FLTKKGQYWT PLNQKALQR GIELFGVGNW KEINYDEFSG KANIVELELR TCMILGINDI TEYYGKKISE EEQEEIKKSN IAKGKKENKL K DNIYQKLQ QMQ

UniProtKB: Transmembrane protein, putative

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Macromolecule #5: S-ADENOSYLMETHIONINE

MacromoleculeName: S-ADENOSYLMETHIONINE / type: ligand / ID: 5 / Number of copies: 1 / Formula: SAM
Molecular weightTheoretical: 398.437 Da
Chemical component information

ChemComp-SAM:
S-ADENOSYLMETHIONINE

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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

Concentration0.25 mg/mL
BufferpH: 8
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 54.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.5 µm / Nominal defocus min: 1.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.6 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 628049
Initial angle assignmentType: PROJECTION MATCHING
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final 3D classificationSoftware - Name: cryoSPARC

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