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- EMDB-62670: Local refinement map of the NTD and MTD regions of BrxX in the Br... -

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Basic information

Entry
Database: EMDB / ID: EMD-62670
TitleLocal refinement map of the NTD and MTD regions of BrxX in the BrxX-Ocr complex
Map data
Sample
  • Complex: The complex of E. coli BrxX methyltransferase with Ocr from bacteriophage T7
Keywordsmethyltransferase / complex / TRANSFERASE
Biological speciesEscherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.18 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
DepositionDec 11, 2024-
Header (metadata) releaseDec 25, 2024-
Map releaseDec 25, 2024-
UpdateDec 25, 2024-
Current statusDec 25, 2024Processing site: PDBc / Status: Released

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Structure visualization

Supplemental images

emd_62670.png

Downloads & links

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Map

FileDownload / File: emd_62670.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

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.86 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.16
Minimum - Maximum-0.26239026 - 0.61483306
Average (Standard dev.)0.00003813408 (±0.008092131)
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: #1

Fileemd_62670_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_62670_half_map_2.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

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

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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
Source (natural)Organism: Escherichia coli (E. coli)

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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

Startup modelType of model: EMDB MAP
EMDB ID:

60037

Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.18 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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