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- EMDB-73869: Cryo-EM structure of Enterotoxigenic Escherichia coli autotranspo... -

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Basic information

Entry
Database: EMDB / ID: EMD-73869
TitleCryo-EM structure of Enterotoxigenic Escherichia coli autotransporter A (EatA) complexed with the fragment antigen binding domain of monoclonal antibody 25
Map datasharpened map
Sample
  • Complex: Binary complex of the recombinant EatA passenger domain with Fab 25
    • Complex: Enterotoxigenic Escherichia coli autotransporter A (EatA) passenger domain
      • Protein or peptide: Serine protease EatA
    • Complex: Fragment antigen binding domain of monoclonal antibody 25
      • Protein or peptide: Heavy chain of the fragment antigen binding domain of monoclonal antibody 25
      • Protein or peptide: Light chain of the fragment antigen binding domain of monoclonal antibody 25
KeywordsPROTEASE / BETA-HELIX / SECRETED / MONOCLONAL ANTIBODY / HYDROLASE / HYDROLASE-IMMUNE SYSTEM complex
Function / homology
Function and homology information


Hydrolases; Acting on peptide bonds (peptidases); Serine endopeptidases / cell outer membrane / periplasmic space / serine-type endopeptidase activity / cell surface / proteolysis / extracellular region
Similarity search - Function
: / PIC/HAP1/IgA0 second beta-solenoid repeat region / : / Peptidase S6, IgA endopeptidase / Peptidase family S6 domain / Immunoglobulin A1 protease / Peptidase family S6 domain profile. / Autotransporter beta-domain / Outer membrane autotransporter barrel / Autotransporter beta-domain ...: / PIC/HAP1/IgA0 second beta-solenoid repeat region / : / Peptidase S6, IgA endopeptidase / Peptidase family S6 domain / Immunoglobulin A1 protease / Peptidase family S6 domain profile. / Autotransporter beta-domain / Outer membrane autotransporter barrel / Autotransporter beta-domain / Autotransporter beta-domain profile. / Autotransporter beta-domain / Autotransporter beta-domain superfamily / Autotransporter, pectate lyase C-like domain superfamily / Pectin lyase fold/virulence factor / Peptidase S1, PA clan
Similarity search - Domain/homology
Serine protease EatA
Similarity search - Component
Biological speciesEscherichia coli (E. coli) / Homo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.32 Å
AuthorsBuckley DP / Berndsen ZT
Funding support United States, 4 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)AI089894 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)AI126887 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)T32AI007172 United States
Department of Veterans Affairs (VA, United States)5I01BX001469-05 United States
CitationJournal: To Be Published
Title: Human infection with enterotoxigenic E. coli elicits antibodies that broadly neutralize mucin-degrading proteases of pathogenic E. coli and Shigella
Authors: Buckley DP / Akhtar M / Thapa M / Schmitz A / Turner J / Vickers TJ / Khatoon N / Kaisar H / Coggin JA / Ganguli D / Sheikh A / Laird RM / Poly F / Porter CK / Ruiz-Perez F / Miller MJ / ...Authors: Buckley DP / Akhtar M / Thapa M / Schmitz A / Turner J / Vickers TJ / Khatoon N / Kaisar H / Coggin JA / Ganguli D / Sheikh A / Laird RM / Poly F / Porter CK / Ruiz-Perez F / Miller MJ / Bhuiyan TR / Qadri F / Trillo-Muyo S / Dolan B / van der Post S / Ellebedy A / Berndsen ZT / Fleckenstein JM
History
DepositionNov 16, 2025-
Header (metadata) releaseJun 3, 2026-
Map releaseJun 3, 2026-
UpdateJun 3, 2026-
Current statusJun 3, 2026Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_73869.png

Downloads & links

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Map

FileDownload / File: emd_73869.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationsharpened map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.93 Å/pix.
x 320 pix.
= 296.96 Å		0.93 Å/pix.
x 320 pix.
= 296.96 Å		0.93 Å/pix.
x 320 pix.
= 296.96 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.928 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.642
Minimum - Maximum-2.2093732 - 3.2237778
Average (Standard dev.)-0.00047927568 (±0.06975905)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 296.96 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: half map A

Fileemd_73869_half_map_1.map
Annotationhalf map A
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half map B

Fileemd_73869_half_map_2.map
Annotationhalf map B
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Binary complex of the recombinant EatA passenger domain with Fab 25

EntireName: Binary complex of the recombinant EatA passenger domain with Fab 25
Components
  • Complex: Binary complex of the recombinant EatA passenger domain with Fab 25
    • Complex: Enterotoxigenic Escherichia coli autotransporter A (EatA) passenger domain
      • Protein or peptide: Serine protease EatA
    • Complex: Fragment antigen binding domain of monoclonal antibody 25
      • Protein or peptide: Heavy chain of the fragment antigen binding domain of monoclonal antibody 25
      • Protein or peptide: Light chain of the fragment antigen binding domain of monoclonal antibody 25

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Supramolecule #1: Binary complex of the recombinant EatA passenger domain with Fab 25

SupramoleculeName: Binary complex of the recombinant EatA passenger domain with Fab 25
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Escherichia coli (E. coli) / Strain: H10407
Molecular weightTheoretical: 23 KDa

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Supramolecule #2: Enterotoxigenic Escherichia coli autotransporter A (EatA) passeng...

SupramoleculeName: Enterotoxigenic Escherichia coli autotransporter A (EatA) passenger domain
type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1
Details: Mature, secreted EatA passenger domain (cleaved N-terminal signal peptide and C-terminal beta-barrel regions) from recombinant expression of full-length EatA
Source (natural)Organism: Escherichia coli (E. coli) / Strain: H10407

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Supramolecule #3: Fragment antigen binding domain of monoclonal antibody 25

SupramoleculeName: Fragment antigen binding domain of monoclonal antibody 25
type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2-#3
Details: Fab fragment generated from papain-digested monoclonal 25 IgG
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Serine protease EatA

MacromoleculeName: Serine protease EatA / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
EC number: Hydrolases; Acting on peptide bonds (peptidases); Serine endopeptidases
Source (natural)Organism: Escherichia coli (E. coli) / Strain: H10407
Molecular weightTheoretical: 111.014391 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: ATVNADISYQ TFRDFAENKG AFIVGASNIN IYDKNGVLVG VLDKAPMPDF SSATMNTGTL PPGDHTLYSP QYVVTAKHVN GSDIMSFGH IQNNYTVVGE NNHNSLDIKI RRLNKIVTEV APAEISSVGA VNGAYQEGGR FKAFYRLGGG LQYIKDKNGN L TPVYTNGG ...String:
ATVNADISYQ TFRDFAENKG AFIVGASNIN IYDKNGVLVG VLDKAPMPDF SSATMNTGTL PPGDHTLYSP QYVVTAKHVN GSDIMSFGH IQNNYTVVGE NNHNSLDIKI RRLNKIVTEV APAEISSVGA VNGAYQEGGR FKAFYRLGGG LQYIKDKNGN L TPVYTNGG FLTGGTISAL SSYNNGQMIT APTGDIFNPA NGPLANYLNK GDSGSPLFAY DSLDKKWVLV GVLSSGSEHG NN WVVTTQD FLHQQPKHDF DKTISYDSEK GSLQWRYNKN SGVGTLSQES VVWDMHGKKG GDLNAGKNLQ FTGNNGEIIL HDS IDQGAG YLQFFDNYTV TSLTDQTWTG GGIITEKGVN VLWQVNGVND DNLHKVGEGT LTVNGKGVNN GGLKVGDGTV ILNQ RPDDN GHKQAFSSIN ISSGRATVIL SDANQVNPDK ISWGYRGGTL DLNGNNVNFT RLQAADYGAI VSNNNKNKSE LTLKL QTLN ENDISVDVKT YEVFGGHGSP GDLYYVPASN TYFILKSKAY GPFFSDLDNT NVWQNVGHDR DKAIQIVKQQ KIGESS QPY MFHGQLNGYM DVNIHPLSGK DVLTLDGSVN LPEGVITKKS GTLIFQGHPV IHAGMTTSAG QSDWENRQFT MDKLRLD AA TFHLSRNAHM QGDISAANGS TVILGSSRVF TDKNDGTGNA VSSVEGSSIA TTAGDQSYYS GNVLLENHSS LEVRENFT G GIEAYDSSVS VTSQNAIFDH VGSFVNSSLL LEKGAKLTAQ SGIFTNNTMK IKENASLTLT GIPSVGKPGY YSPVTSTTE GIHLGERASL SVKNMGYLSS NITAENSAAI INLGDSNATI GKTDSPLFST LMRGYNAVLQ GNIMGPQSSV NMNNALWHSD RNSELKELK ANDSQIELGV RGHFAKLRVK ELIASNSVFL VHANNSQADQ LNVTDKLQGS NNTILVDFFN KAANGTNVTL I TAPKGSDE NTFKAGTQQI GFSNITPEIR TENTDTATQW VLTGYQSVAD ARASKIATDF MDSGYKSFLT EVN

UniProtKB: Serine protease EatA

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Macromolecule #2: Heavy chain of the fragment antigen binding domain of monoclonal ...

MacromoleculeName: Heavy chain of the fragment antigen binding domain of monoclonal antibody 25
type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 24.135098 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: QVQLQESGPG LVKPSQTLSL TCTVSGGSIT SGDYYWSWIR QPPGKGLEWI GHISHSGTMY YNPSLKSRLS ISVDTSKNQF SLNLSSVTA ADTAVYYCAR GGFSMLRGVP SVLAYWGQGT LVTVSSASTK GPSVFPLAPS SKSTSGGTAA LGCLVKDYFP E PVTVSWNS ...String:
QVQLQESGPG LVKPSQTLSL TCTVSGGSIT SGDYYWSWIR QPPGKGLEWI GHISHSGTMY YNPSLKSRLS ISVDTSKNQF SLNLSSVTA ADTAVYYCAR GGFSMLRGVP SVLAYWGQGT LVTVSSASTK GPSVFPLAPS SKSTSGGTAA LGCLVKDYFP E PVTVSWNS GALTSGVHTF PAVLQSSGLY SLSSVVTVPS SSLGTQTYIC NVNHKPSNTK VDKRVEPKSC D

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Macromolecule #3: Light chain of the fragment antigen binding domain of monoclonal ...

MacromoleculeName: Light chain of the fragment antigen binding domain of monoclonal antibody 25
type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 23.272832 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: DIQMTQSPSS LSASVGDRVT ITCRASESIS SYLNWYQQKP GKAPKLLIYA ASSLQSGVPS RFSGRGSGTD FTLTISSLQP EDFATYHCQ QSFTTPLTFG GGTKVEIKRT VAAPSVFIFP PSDEQLKSGT ASVVCLLNNF YPREAKVQWK VDNALQSGNS Q ESVTEQDS ...String:
DIQMTQSPSS LSASVGDRVT ITCRASESIS SYLNWYQQKP GKAPKLLIYA ASSLQSGVPS RFSGRGSGTD FTLTISSLQP EDFATYHCQ QSFTTPLTFG GGTKVEIKRT VAAPSVFIFP PSDEQLKSGT ASVVCLLNNF YPREAKVQWK VDNALQSGNS Q ESVTEQDS KDSTYSLSST LTLSKADYEK HKVYACEVTH QGLSSPVTKS FNRGEC

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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

Concentration0.7 mg/mL
BufferpH: 7.4 / Component - Concentration: 1.0 X / Component - Formula: TBS / Component - Name: Tris-Buffered Saline
Details: 10X TBS: 250mM Tris, 27mM potassium chloride, 1.37M sodium chloride, pH 7.4
GridModel: UltrAuFoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: GOLD / Support film - topology: HOLEY / Support film - Film thickness: 50 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 15 sec. / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 0.039 kPa
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV
Details: Added 1X Lauryl Maltose Neopentyl Glycol (LMNG) detergent to sample prior to vitrification..
DetailsThis sample was monodisperse.

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Electron microscopy

MicroscopeTFS GLACIOS
SoftwareName: EPU / Details: TFS EPU
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Number grids imaged: 1 / Number real images: 4748 / Average exposure time: 8.72 sec. / Average electron dose: 54.0 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.2 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 150000
Sample stageCooling holder cryogen: NITROGEN

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Image processing

Particle selectionNumber selected: 1272593
Details: Initial particle set was obtained by CryoSegNet AI picking, followed by cryoSPARC 2D classification to redo particle picking with 2D templates
CTF correctionSoftware - Name: cryoSPARC (ver. 4.7.1) / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: OTHER / Details: AB-INITIO
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.32 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.7.1) / Number images used: 33847
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.7.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.7.1)
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelChain - Source name: AlphaFold / Chain - Initial model type: in silico model
SoftwareName: UCSF ChimeraX (ver. 1.8)
DetailsRigid body fitting of AlphaFold-derived models was done in ChimeraX, followed by multiple rounds of local fitting in Coot and Phenix real-space refinement
RefinementSpace: REAL / Protocol: FLEXIBLE FIT / Target criteria: Cross-correlation coefficient
Output model

PDB-9z76:
Cryo-EM structure of Enterotoxigenic Escherichia coli autotransporter A (EatA) complexed with the fragment antigen binding domain of monoclonal antibody 25

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