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- EMDB-16807: Cryo-EM structure of PcrV/Fab(11-E5) -

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Basic information

Entry
Database: EMDB / ID: EMD-16807
TitleCryo-EM structure of PcrV/Fab(11-E5)
Map data
Sample
  • Complex: PcrV-Fab(11-E5)
    • Protein or peptide: Heavy chain
    • Protein or peptide: Light chain
    • Protein or peptide: Maltose/maltodextrin-binding periplasmic protein,Type III secretion protein PcrV
Keywordsantibody T3SS Tip protein / ANTIMICROBIAL PROTEIN
Function / homology
Function and homology information


type III protein secretion system complex / protein secretion by the type III secretion system / detection of maltose stimulus / maltose transport complex / carbohydrate transport / carbohydrate transmembrane transporter activity / maltose binding / maltose transport / maltodextrin transmembrane transport / ATP-binding cassette (ABC) transporter complex, substrate-binding subunit-containing ...type III protein secretion system complex / protein secretion by the type III secretion system / detection of maltose stimulus / maltose transport complex / carbohydrate transport / carbohydrate transmembrane transporter activity / maltose binding / maltose transport / maltodextrin transmembrane transport / ATP-binding cassette (ABC) transporter complex, substrate-binding subunit-containing / ATP-binding cassette (ABC) transporter complex / cell chemotaxis / outer membrane-bounded periplasmic space / periplasmic space / DNA damage response / extracellular space / membrane
Similarity search - Function
Low calcium response V antigen / Virulence-associated V antigen superfamily / V antigen (LcrV) protein / Maltose/Cyclodextrin ABC transporter, substrate-binding protein / Solute-binding family 1, conserved site / Bacterial extracellular solute-binding proteins, family 1 signature. / Bacterial extracellular solute-binding protein / Bacterial extracellular solute-binding protein
Similarity search - Domain/homology
Type III secretion protein PcrV / Maltose/maltodextrin-binding periplasmic protein
Similarity search - Component
Biological speciesHomo sapiens (human) / Pseudomonas aeruginosa (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.6 Å
AuthorsYuan B / Simonis A / Marlovits TC
Funding support Germany, 1 items
OrganizationGrant numberCountry
German Research Foundation (DFG) Germany
CitationJournal: Cell / Year: 2023
Title: Discovery of highly neutralizing human antibodies targeting Pseudomonas aeruginosa.
Authors: Alexander Simonis / Christoph Kreer / Alexandra Albus / Katharina Rox / Biao Yuan / Dmitriy Holzmann / Joana A Wilms / Sylvia Zuber / Lisa Kottege / Sandra Winter / Meike Meyer / Kristin ...Authors: Alexander Simonis / Christoph Kreer / Alexandra Albus / Katharina Rox / Biao Yuan / Dmitriy Holzmann / Joana A Wilms / Sylvia Zuber / Lisa Kottege / Sandra Winter / Meike Meyer / Kristin Schmitt / Henning Gruell / Sebastian J Theobald / Anna-Maria Hellmann / Christina Meyer / Meryem Seda Ercanoglu / Nina Cramer / Antje Munder / Michael Hallek / Gerd Fätkenheuer / Manuel Koch / Harald Seifert / Ernst Rietschel / Thomas C Marlovits / Silke van Koningsbruggen-Rietschel / Florian Klein / Jan Rybniker /
Abstract: Drug-resistant Pseudomonas aeruginosa (PA) poses an emerging threat to human health with urgent need for alternative therapeutic approaches. Here, we deciphered the B cell and antibody response to ...Drug-resistant Pseudomonas aeruginosa (PA) poses an emerging threat to human health with urgent need for alternative therapeutic approaches. Here, we deciphered the B cell and antibody response to the virulence-associated type III secretion system (T3SS) in a cohort of patients chronically infected with PA. Single-cell analytics revealed a diverse B cell receptor repertoire directed against the T3SS needle-tip protein PcrV, enabling the production of monoclonal antibodies (mAbs) abrogating T3SS-mediated cytotoxicity. Mechanistic studies involving cryoelectron microscopy identified a surface-exposed C-terminal PcrV epitope as the target of highly neutralizing mAbs with broad activity against drug-resistant PA isolates. These anti-PcrV mAbs were as effective as treatment with conventional antibiotics in vivo. Our study reveals that chronically infected patients represent a source of neutralizing antibodies, which can be exploited as therapeutics against PA.
History
DepositionMar 8, 2023-
Header (metadata) releaseNov 22, 2023-
Map releaseNov 22, 2023-
UpdateNov 22, 2023-
Current statusNov 22, 2023Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_16807.png

Downloads & links

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Map

FileDownload / File: emd_16807.map.gz / Format: CCP4 / Size: 18.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.85 Å/pix.
x 168 pix.
= 142.8 Å		0.85 Å/pix.
x 168 pix.
= 142.8 Å		0.85 Å/pix.
x 168 pix.
= 142.8 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.85 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.00418
Minimum - Maximum-0.018828439 - 0.026291931
Average (Standard dev.)0.00018123873 (±0.001376246)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions168168168
Spacing168168168
CellA=B=C: 142.8 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_16807_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_16807_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : PcrV-Fab(11-E5)

EntireName: PcrV-Fab(11-E5)
Components
  • Complex: PcrV-Fab(11-E5)
    • Protein or peptide: Heavy chain
    • Protein or peptide: Light chain
    • Protein or peptide: Maltose/maltodextrin-binding periplasmic protein,Type III secretion protein PcrV

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Supramolecule #1: PcrV-Fab(11-E5)

SupramoleculeName: PcrV-Fab(11-E5) / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Heavy chain

MacromoleculeName: Heavy chain / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 24.036674 KDa
SequenceString: EVQLLESGGG LVQPGGSLRL SCAASGFSFS SYAMSWVRQA PGKGLEWVSA ISGSGGITYY GDSAKGRFTI SRDNSKNTLY LEMSSLRAD DTAVYYCAQE RYCDSGSCYE RDPVFEYWGQ GTRVTVSSAS TKGPSVFPLA PSSKSTSGGT AALGCLVKDY F PEPVTVSW ...String:
EVQLLESGGG LVQPGGSLRL SCAASGFSFS SYAMSWVRQA PGKGLEWVSA ISGSGGITYY GDSAKGRFTI SRDNSKNTLY LEMSSLRAD DTAVYYCAQE RYCDSGSCYE RDPVFEYWGQ GTRVTVSSAS TKGPSVFPLA PSSKSTSGGT AALGCLVKDY F PEPVTVSW NSGALTSGVH TFPAVLQSSG LYSLSSVVTV PSSSLGTQTY ICNVNHKPSN TKVDKRVEP

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Macromolecule #2: Light chain

MacromoleculeName: Light chain / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 22.964422 KDa
SequenceString: QSVLTQPPSA SGAPGQRVTI SCSGSNSNIG TYFVYWYQQL PGTAPKVLIY RNDQRPSGVP DRISGSKSGT SASLAISGLR SEDEADYYC ASWDASLRGY VFGPGTKVTV LGQPKAAPSV TLFPPSSEEL QANKATLVCL ISDFYPGAVT VAWKADSSPV K AGVETTTP ...String:
QSVLTQPPSA SGAPGQRVTI SCSGSNSNIG TYFVYWYQQL PGTAPKVLIY RNDQRPSGVP DRISGSKSGT SASLAISGLR SEDEADYYC ASWDASLRGY VFGPGTKVTV LGQPKAAPSV TLFPPSSEEL QANKATLVCL ISDFYPGAVT VAWKADSSPV K AGVETTTP SKQSNNKYAA SSYLSLTPEQ WKSHRSYSCQ VTHEGSTVEK TVAPTECS

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Macromolecule #3: Maltose/maltodextrin-binding periplasmic protein,Type III secreti...

MacromoleculeName: Maltose/maltodextrin-binding periplasmic protein,Type III secretion protein PcrV
type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Pseudomonas aeruginosa (bacteria)
Molecular weightTheoretical: 75.538055 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MGSWSHPQFE KGGGSGGGSG GGSWSHPQFE KSGLVPRGSA SKTEEGKLVI WINGDKGYNG LAEVGKKFEK DTGIKVTVEH PDKLEEKFP QVAATGDGPD IIFWAHDRFG GYAQSGLLAE ITPAAAFQDK LYPFTWDAVR YNGKLIAYPI AVEALSLIYN K DLLPNPPK ...String:
MGSWSHPQFE KGGGSGGGSG GGSWSHPQFE KSGLVPRGSA SKTEEGKLVI WINGDKGYNG LAEVGKKFEK DTGIKVTVEH PDKLEEKFP QVAATGDGPD IIFWAHDRFG GYAQSGLLAE ITPAAAFQDK LYPFTWDAVR YNGKLIAYPI AVEALSLIYN K DLLPNPPK TWEEIPALDK ELKAKGKSAL MFNLQEPYFT WPLIAADGGY AFKYAAGKYD IKDVGVDNAG AKAGLTFLVD LI KNKHMNA DTDYSIAEHA FNHGETAMTI NGPWAWSNID TSKVNYGVTV LPTFKGQPSK PFVGVLSAGI NAASPNKELA KEF LENYLL TDEGLEAVNK DKPLGAVALK SYEEELVKDP RVAATMENAQ KGEIMPNIPQ MSAFWYAVRT AVINAASGRQ TVDE VRNLN AARELFLDEL LAAPAAPASA EQEELLALLR SERIVLAHAG QPLSEAQVLK ALAWLLAANP SAPPGQGLEV LREVL QARR QPGAQWDLRE FLVSAYFSLH GRLDEDVIGV YKDVLQTQDG KRKALLDELK ALTAELKVYS VIQSQINAAL SAKQGI RID AGGIDLVDPT LYGYAVGDPR WKDSPEYALL SNLDTFSGKL SIKDFLSGSP KQSGELKGLK DEYPFEKDNN PVGNFAT TV SDRSRPLNDK VNEKTTLLND TSSRYNSAVE ALNRFIQKYD SVLRDILSAI

UniProtKB: Maltose/maltodextrin-binding periplasmic protein, Type III secretion protein PcrV

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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

Concentration0.33 mg/mL
BufferpH: 7.4 / Details: 1xPBS
VitrificationCryogen name: ETHANE-PROPANE / Chamber humidity: 100 %

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 1.24 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER
Final reconstructionResolution.type: BY AUTHOR / Resolution: 4.6 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 99625
Initial angle assignmentType: ANGULAR RECONSTITUTION
Final angle assignmentType: ANGULAR RECONSTITUTION
FSC plot (resolution estimation)

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