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- EMDB-43613: Structure of HamAB apo complex from the Escherichia coli Hachiman... -

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Basic information

Entry
Database: EMDB / ID: EMD-43613
TitleStructure of HamAB apo complex from the Escherichia coli Hachiman defense system
Map dataSharp map of the apo HamAB complex
Sample
  • Complex: Apo HamAB complex
    • Protein or peptide: HamA
    • Protein or peptide: HamB
KeywordsNuclease-helicase complex / antiphage defense system / IMMUNE SYSTEM
Function / homology
Function and homology information


helicase activity / nucleic acid binding / hydrolase activity / ATP binding
Similarity search - Function
Anti-bacteriophage protein A/HamA, C-terminal domain / HamA / DEAD/DEAH box helicase / DEAD/DEAH box helicase domain / 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 / DUF1837 domain-containing protein
Similarity search - Component
Biological speciesEscherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.65 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 3, 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_43613.png

Downloads & links

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Map

FileDownload / File: emd_43613.map.gz / Format: CCP4 / Size: 115.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSharp map of the apo HamAB complex
Voxel sizeX=Y=Z: 1.048 Å
Density
Contour LevelBy AUTHOR: 0.05
Minimum - Maximum-0.001788998 - 2.1177518
Average (Standard dev.)0.0005685334 (±0.01682685)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions312312312
Spacing312312312
CellA=B=C: 326.97598 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_43613_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 the apo HamAB complex

Fileemd_43613_additional_1.map
AnnotationUnsharpened map of the apo HamAB complex
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map A of the apo HamAB complex

Fileemd_43613_half_map_1.map
AnnotationHalf map A of the apo HamAB complex
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map B of the apo HamAB complex

Fileemd_43613_half_map_2.map
AnnotationHalf map B of the apo HamAB 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 : Apo HamAB complex

EntireName: Apo HamAB complex
Components
  • Complex: Apo HamAB complex
    • Protein or peptide: HamA
    • Protein or peptide: HamB

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Supramolecule #1: Apo HamAB complex

SupramoleculeName: Apo HamAB complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Escherichia coli (E. coli) / Strain: ECOR31
Molecular weightTheoretical: 160 KDa

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Macromolecule #1: HamA

MacromoleculeName: HamA / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Escherichia coli (E. coli) / Strain: ECOR31
Molecular weightTheoretical: 28.923797 KDa
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria)
SequenceString: MANFEDWCDS TERNISDHYL QSITARDAEC MFGVQVMAAL IPEHYASPRN IANAFEALGK PGLAAYIAGK LPETKQIRSG DLGEIFATE WINARSNGYK TPIKRLRWKD HRNMSMRGED VIGIYIDQSS QQLFFLKTEA KSRAKMTGEV VSEARDNLNK E QGLPSSHA ...String:
MANFEDWCDS TERNISDHYL QSITARDAEC MFGVQVMAAL IPEHYASPRN IANAFEALGK PGLAAYIAGK LPETKQIRSG DLGEIFATE WINARSNGYK TPIKRLRWKD HRNMSMRGED VIGIYIDQSS QQLFFLKTEA KSRAKMTGEV VSEARDNLNK E QGLPSSHA LMFIADRLNE QGEELLAKAI LNATLRQGIV PGCVRHLIFL LSGNSSETML TTSIEKYTGQ NNQWGVCLRI AR HGEFIAA TFEKVISDAS NS

UniProtKB: DUF1837 domain-containing protein

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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 BL21 (bacteria)
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 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
20.0 mMC8H18N2O4SHEPES
100.0 mMKClpotassium chloride
1.0 mMC9H15O6PTCEP
0.5 %C3H8O3glycerol
GridModel: UltrAuFoil R2/2 / Material: GOLD / Mesh: 200 / Support film - Material: GOLD / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 15 sec. / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 39.0 kPa
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 281 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: OTHER / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 1.8 µm / Nominal defocus min: 0.8 µm
Sample stageCooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number grids imaged: 1 / Number real images: 4796 / Average electron dose: 50.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 5538550
Startup modelType of model: OTHER / Details: cryoSPARC ab initio model
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.2.1)
Final 3D classificationSoftware - Name: cryoSPARC (ver. 4.2.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.2.1)
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.65 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.2.1) / Number images used: 309630
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-8vx9:
Structure of HamAB apo complex from the Escherichia coli Hachiman defense system

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