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- EMDB-60037: Cryo-EM structure of the E. coli BrxX methyltransferase complexed... -

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Basic information

Entry
Database: EMDB / ID: EMD-60037
TitleCryo-EM structure of the E. coli BrxX methyltransferase complexed with Ocr
Map data
Sample
  • Complex: The complex of E. coli BrxX methyltransferase with Ocr from bacteriophage T7
    • Protein or peptide: site-specific DNA-methyltransferase (adenine-specific)
    • Protein or peptide: Protein Ocr
Keywordsmethyltransferase / complex / TRANSFERASE
Function / homology
Function and homology information


symbiont-mediated evasion of host restriction-modification system / DNA modification / site-specific DNA-methyltransferase (adenine-specific) / site-specific DNA-methyltransferase (adenine-specific) activity / methylation / nucleic acid binding / symbiont-mediated suppression of host innate immune response
Similarity search - Function
: / B-form DNA mimic Ocr / DNA mimic ocr / Protein Ocr / Type II restriction enzyme and methyltransferase RM.Eco57I-like / Eco57I restriction-modification methylase / : / N-6 Adenine-specific DNA methylases signature. / DNA methylase, N-6 adenine-specific, conserved site / S-adenosyl-L-methionine-dependent methyltransferase superfamily
Similarity search - Domain/homology
site-specific DNA-methyltransferase (adenine-specific) / Protein Ocr
Similarity search - Component
Biological speciesEscherichia coli (E. coli) / Escherichia phage T7 (virus)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.15 Å
AuthorsZhu L / Xu TH / Sun LT
Funding support China, 2 items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)32271314 China
National Natural Science Foundation of China (NSFC)31600593 China
CitationJournal: Nucleic Acids Res / Year: 2024
Title: Ocr-mediated suppression of BrxX unveils a phage counter-defense mechanism.
Authors: Shen Li / Tianhao Xu / Xinru Meng / Yujuan Yan / Ying Zhou / Lei Duan / Yulong Tang / Li Zhu / Litao Sun /
Abstract: The burgeoning crisis of antibiotic resistance has directed attention to bacteriophages as natural antibacterial agents capable of circumventing bacterial defenses. Central to this are the bacterial ...The burgeoning crisis of antibiotic resistance has directed attention to bacteriophages as natural antibacterial agents capable of circumventing bacterial defenses. Central to this are the bacterial defense mechanisms, such as the BREX system, which utilizes the methyltransferase BrxX to protect against phage infection. This study presents the first in vitro characterization of BrxX from Escherichia coli, revealing its substrate-specific recognition and catalytic activity. We demonstrate that BrxX exhibits nonspecific DNA binding but selectively methylates adenine within specific motifs. Kinetic analysis indicates a potential regulation of BrxX by the concentration of its co-substrate, S-adenosylmethionine, and suggests a role for other BREX components in modulating BrxX activity. Furthermore, we elucidate the molecular mechanism by which the T7 phage protein Ocr (Overcoming classical restriction) inhibits BrxX. Despite low sequence homology between BrxX from different bacterial species, Ocr effectively suppresses BrxX's enzymatic activity through high-affinity binding. Cryo-electron microscopy and biophysical analyses reveal that Ocr, a DNA mimic, forms a stable complex with BrxX, highlighting a conserved interaction interface across diverse BrxX variants. Our findings provide insights into the strategic counteraction by phages against bacterial defense systems and offer a foundational understanding of the complex interplay between phages and their bacterial hosts, with implications for the development of phage therapy to combat antibiotic resistance.
History
DepositionMay 6, 2024-
Header (metadata) releaseDec 11, 2024-
Map releaseDec 11, 2024-
UpdateJul 16, 2025-
Current statusJul 16, 2025Processing site: PDBc / Status: Released

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Structure visualization

Supplemental images

emd_60037.png

Downloads & links

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Map

FileDownload / File: emd_60037.map.gz / Format: CCP4 / Size: 421.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.86 Å/pix.
x 480 pix.
= 412.8 Å		0.86 Å/pix.
x 480 pix.
= 412.8 Å		0.86 Å/pix.
x 480 pix.
= 412.8 Å

Surface

Projections

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.86 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.1
Minimum - Maximum-0.18722077 - 0.57299167
Average (Standard dev.)0.00004334798 (±0.008771785)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions480480480
Spacing480480480
CellA=B=C: 412.80002 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_60037_half_map_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_60037_half_map_2.map
Projections & Slices
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Histogram (log scale)

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Sample components

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Entire : The complex of E. coli BrxX methyltransferase with Ocr from bacte...

EntireName: The complex of E. coli BrxX methyltransferase with Ocr from bacteriophage T7
Components
  • Complex: The complex of E. coli BrxX methyltransferase with Ocr from bacteriophage T7
    • Protein or peptide: site-specific DNA-methyltransferase (adenine-specific)
    • Protein or peptide: Protein Ocr

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Supramolecule #1: The complex of E. coli BrxX methyltransferase with Ocr from bacte...

SupramoleculeName: The complex of E. coli BrxX methyltransferase with Ocr from bacteriophage T7
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Escherichia coli (E. coli)

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Macromolecule #1: site-specific DNA-methyltransferase (adenine-specific)

MacromoleculeName: site-specific DNA-methyltransferase (adenine-specific)
type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
EC number: site-specific DNA-methyltransferase (adenine-specific)
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 138.043219 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MNTNNIKKYA PQARNDFRDA VIQKLTTLGI AADKKGNLQI AEAETIGETV RYGQFDYPLS TLPRRERLVK RAREQGFEVL VEHCAYTWF NRLCAIRYME LHGYLDHGFR MLSHPETPTA FEVLDHVPEV AEALLPESKA QLVEMKLSGN QDEALYRELL L GQCHALHH ...String:
MNTNNIKKYA PQARNDFRDA VIQKLTTLGI AADKKGNLQI AEAETIGETV RYGQFDYPLS TLPRRERLVK RAREQGFEVL VEHCAYTWF NRLCAIRYME LHGYLDHGFR MLSHPETPTA FEVLDHVPEV AEALLPESKA QLVEMKLSGN QDEALYRELL L GQCHALHH AMPFLFEAVD DEAELLLPDN LTRTDSILRG LVDDIPEEDW EQVEVIGWLY QFYISEKKDA VIGKVVKSED IP AATQLFT PNWIVQYLVQ NSVGRQWLQT YPDSPLKDKM EYYIEPAEQT PEVQAQLAAI TPASIEPESI KVLDPACGSG HIL TEAYNV LKAIYEERGY RTRDIPQLIL ENNIFGLDID DRAAQLSGFA MLMLARQDDR RILGRGVRLN IVSLQESKLD IAEV WTKLN FHQHMQRGSM GDMFTQGTAL ANTDSAEYKL LMRTLALFTS AKTLGSLIQV PQEDEAALKA FLEGLYRLAV EGDIQ QKEA AAELIPYIQQ AWILAQRYDA VVANPPYMGG KGMNGDLKEF AKKQFPDSKS DLFAMFMQHA FSLLKENGFN AQVNMQ SWM FLSSYEALRG WLLDNKTFIT MAHLGARAFG QISGEVVQTT AWVIKNNHSG FYKPVFFRLV DDNEEHKKNN LLNRMNC FK NTLQNDFKKI PGSPIAYWAT LAFINSFLKL PALGTRAVKG LDTNGSIDVF LRRWPEVSIN SFDALGKGNS KWFPIAKG G ELRKWFGNHE YIINYENDGI ELRKNKANLR NKDMYFQEGG TWTVVSTTGF SMRYMPKGFL FDQGGSAVFC ENNDELSIY NILACMNSKY INYSASLICP TLNFTTGDVR KFPVIKNNHL EDLAKKAIEI SKADWNQFET SWEFSKNKLI EHKGNVAYSY ASYCNFQDK LYEQLVNIEK NINNIIEEIL GFKIETTENS ELITLNSNKI YRYGQSETND TFLNRHRSDT ISELISYSVG C QMGRYSLD REGLVYAHEG NKGFADLVAE GAYKTFPADS DGILPLMDDE WFEDDVTSRV KEFVRTVWGE EHLQENLEFI AE SLCLYAI KPKKGESALE TIRRYLSTQF WKDHMKMYKK RPIYWLFSSG KEKAFECLVY LHRYNDATLS RMRTEYVVPL LAR YQANID RLNDQLDEAS GGEATRLKRE RDSLIKKFSE LRSYDDRLRH YADMRISIDL DDGVKVNYGK FGDLLADVKA ITGN APEVI

UniProtKB: site-specific DNA-methyltransferase (adenine-specific)

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Macromolecule #2: Protein Ocr

MacromoleculeName: Protein Ocr / type: protein_or_peptide / ID: 2 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Escherichia phage T7 (virus)
Molecular weightTheoretical: 13.819015 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MAMSNMTYNN VFDHAYEMLK ENIRYDDIRD TDDLHDAIHM AADNAVPHYY ADIFSVMASE GIDLEFEDSG LMPDTKDVIR ILQARIYEQ LTIDLWEDAE DLLNEYLEEV EEYEEDEE

UniProtKB: Protein Ocr

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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

BufferpH: 7.5
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (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: 3.0 µm / Nominal defocus min: 0.9 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.15 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 448285
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementProtocol: RIGID BODY FIT
Output model

PDB-8zek:
Cryo-EM structure of the E. coli BrxX methyltransferase complexed with Ocr

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