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- EMDB-43616: Structure of HamB-DNA complex, conformation 2, from the Escherich... -

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Basic information

Entry
Database: EMDB / ID: EMD-43616
TitleStructure of HamB-DNA complex, conformation 2, from the Escherichia coli Hachiman defense system
Map dataSharp map of conformation 2 of the HamB-DNA complex
Sample
  • Complex: HamB-DNA complex
    • DNA: DNA (40-MER)
    • Protein or peptide: HamB
KeywordsHelicase-DNA complex / antiphage defense system / IMMUNE SYSTEM / IMMUNE SYSTEM-DNA complex
Function / homology
Function and homology information


helicase activity / nucleic acid binding / ATP binding
Similarity search - Function
: / DEAD/DEAH box helicase domain / DEAD/DEAH box helicase / Helicase conserved C-terminal domain / helicase superfamily c-terminal domain / Superfamilies 1 and 2 helicase ATP-binding type-1 domain profile. / DEAD-like helicases superfamily / Helicase, C-terminal / Helicase superfamily 1/2, ATP-binding domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
DEAD/DEAH box helicase
Similarity search - Component
Biological speciesEscherichia coli (E. coli) / synthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.93 Å
AuthorsTuck OT / Doudna JA
Funding support United States, 1 items
OrganizationGrant numberCountry
National Science Foundation (NSF, United States) United States
CitationJournal: bioRxiv / Year: 2024
Title: Hachiman is a genome integrity sensor.
Authors: Owen T Tuck / Benjamin A Adler / Emily G Armbruster / Arushi Lahiri / Jason J Hu / Julia Zhou / Joe Pogliano / Jennifer A Doudna /
Abstract: Hachiman is a broad-spectrum antiphage defense system of unknown function. We show here that Hachiman comprises a heterodimeric nuclease-helicase complex, HamAB. HamA, previously a protein of unknown ...Hachiman is a broad-spectrum antiphage defense system of unknown function. We show here that Hachiman comprises a heterodimeric nuclease-helicase complex, HamAB. HamA, previously a protein of unknown function, is the effector nuclease. HamB is the sensor helicase. HamB constrains HamA activity during surveillance of intact dsDNA. When the HamAB complex detects DNA damage, HamB helicase activity liberates HamA, unleashing nuclease activity. Hachiman activation degrades all DNA in the cell, creating 'phantom' cells devoid of both phage and host DNA. We demonstrate Hachiman activation in the absence of phage by treatment with DNA-damaging agents, suggesting that Hachiman responds to aberrant DNA states. Phylogenetic similarities between the Hachiman helicase and eukaryotic enzymes suggest this bacterial immune system has been repurposed for diverse functions across all domains of life.
History
DepositionFeb 4, 2024-
Header (metadata) releaseMar 13, 2024-
Map releaseMar 13, 2024-
UpdateMar 20, 2024-
Current statusMar 20, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_43616.png

Downloads & links

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Map

FileDownload / File: emd_43616.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSharp map of conformation 2 of the HamB-DNA complex
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.12 Å/pix.
x 256 pix.
= 285.44 Å		1.12 Å/pix.
x 256 pix.
= 285.44 Å		1.12 Å/pix.
x 256 pix.
= 285.44 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.115 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.075
Minimum - Maximum-0.0017294913 - 2.0530934
Average (Standard dev.)0.0008136213 (±0.020492159)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 285.44 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_43616_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Unsharpened map of conformation 2 of the HamB-DNA complex

Fileemd_43616_additional_1.map
AnnotationUnsharpened map of conformation 2 of the HamB-DNA complex
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map A of conformation 2 of the HamB-DNA complex

Fileemd_43616_half_map_1.map
AnnotationHalf map A of conformation 2 of the HamB-DNA complex
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map B of conformation 2 of the HamB-DNA complex

Fileemd_43616_half_map_2.map
AnnotationHalf map B of conformation 2 of the HamB-DNA 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 : HamB-DNA complex

EntireName: HamB-DNA complex
Components
  • Complex: HamB-DNA complex
    • DNA: DNA (40-MER)
    • Protein or peptide: HamB

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Supramolecule #1: HamB-DNA complex

SupramoleculeName: HamB-DNA complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Escherichia coli (E. coli) / Strain: ECOR31
Molecular weightTheoretical: 155 KDa

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Macromolecule #1: DNA (40-MER)

MacromoleculeName: DNA (40-MER) / type: dna / ID: 1 / Number of copies: 2 / Classification: DNA
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 12.154847 KDa
SequenceString:
(DT)(DC)(DG)(DT)(DC)(DA)(DC)(DC)(DA)(DG) (DT)(DA)(DC)(DA)(DA)(DA)(DC)(DT)(DA)(DC) (DA)(DA)(DC)(DG)(DC)(DC)(DT)(DG)(DT) (DA)(DG)(DC)(DA)(DT)(DT)(DC)(DC)(DA)(DC) (DA)

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Macromolecule #2: HamB

MacromoleculeName: HamB / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Escherichia coli (E. coli) / Strain: ECOR31
Molecular weightTheoretical: 130.898453 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MPATADEIIE AIKEASAVGF RGRLIARGQA RSVIWRDGDL PPDAPEFSAL LSQDLQGYAY ALIDLGLRLR ELNGDDAYAR IAFEQAGTA LESAIAKGKR DSRDTDFHFV MAAASYHLAH LSARAYSLLA MVGQDDNFSP IERALTQLIR RDLRTLRDNA L GFRLRGDG ...String:
MPATADEIIE AIKEASAVGF RGRLIARGQA RSVIWRDGDL PPDAPEFSAL LSQDLQGYAY ALIDLGLRLR ELNGDDAYAR IAFEQAGTA LESAIAKGKR DSRDTDFHFV MAAASYHLAH LSARAYSLLA MVGQDDNFSP IERALTQLIR RDLRTLRDNA L GFRLRGDG SDVKITEILQ ARLNLPQDEN GDSESEEDIL FDGLDLALTD AYMSAISLYL LAVERGESRL LSRAIEKLRI SL SICAQFN MLPQWWLNFI TIHLLSDLWS DTFHERLPLV PVGGDAAEWP ALRELFIALL QRRPRAEIDL WPSQREAAGR SVN DNDDLV VSLPTSAGKT RIAELCILRC LAGGKRVVFI TPLRALSAQT EATLSRTFGP LGKTISMLYG SIGVSGMDED AIRQ RDIVV ATPEKLDFAL RNDPSIINDV GLFIFDEGHM IGADEREVRY EVQIQRLLRR QDADTRRIVC LSAILPDGEQ LDDFA GWLR RDKPGGPIKN NWRPTRLQFG EVIWSAPAGR LNLSVGYEAA WVSRFIVSRQ PPKVKLPNKK QRTKMFPSDN KELCLA TAW RLIEDGQTVL IYCPLRRSVE PFAETIVDLH QRGLLPSLFD AAPDILDTAI SLGEEWLGAH SPILACLRLG VALHHGA LP TAYRKEIERL LRDGVLKVTI SSPTLAQGLN LSATAIVMHS LHRNRELIKV SEFRNVIGRA GRAYVDVEGL VIYPIFDK V NKRQTNWHTL TSDTGAREME SGLIQLVCVL LIRMHTRLGG DLKALTEYVT NNAVAWEFPE IMTESPQERD IAQAIWEKQ LSTLDTAILS LLGENDIPDD QIETALDDIL QSSLWQRSLQ RYRDENERIL LKSGLLSRSR YIWQRSTAAG RRGYFLSGVG LTTGLRLDA IAAKANQLLI DANAAIMGGD AEEAIAAITA LAEEVFTFYP FIPDPLPGDW RGILRSWLLG EPMTNVANTQ A SETLQFVE NGLVYRLPWA MEAIRVRATA NGDLIGDTDT TLDDYELGFA VAAVETGTLS RSSSLLIQAG FSSRLAAIKV VT DTTADFQ SGQELRRWLN SEEVISHTDN HDWPTPETRV MWLEFLGSLS PKGSQVWSRH RYNGMVDWRD TPAVIGTPLQ LYT VDGIHH VLADDGTPLG SINGRINTNR RGLLRVEVDD ENGRAMFDYL GPDDFIST

UniProtKB: DEAD/DEAH box helicase

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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

Concentration1.5 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
20.0 mMC8H18N2O4SHEPES
100.0 mMKClpotassium chloride
1.0 mMC9H15O6PTCEP
0.5 %C3H8O3glycerol
2.0 mMMgCl2magnesium chloride
1.0 mMC10H16N5O13P3adenosine triphosphate
GridModel: UltrAuFoil R2/2 / Material: GOLD / Support film - Material: GOLD / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 281 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TALOS ARCTICA
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number grids imaged: 1 / Number real images: 9133 / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 1.8 µm / Nominal defocus min: 0.8 µm
Sample stageCooling holder cryogen: NITROGEN
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 18254957
Startup modelType of model: OTHER / Details: cryoSPARC ab initio model
Final reconstructionAlgorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 2.93 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.2.1) / Number images used: 249313
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.2.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.2.1)
Final 3D classificationSoftware - Name: cryoSPARC (ver. 4.2.1)
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelChain - Source name: AlphaFold / Chain - Initial model type: in silico model / Details: v1.4.0
RefinementSpace: REAL / Protocol: OTHER
Output model

PDB-8vxc:
Structure of HamB-DNA complex, conformation 2, from the Escherichia coli Hachiman defense system

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