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- PDB-8p2v: Neisseria meningitidis Type IV pilus SB-GATDH variant -

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Basic information

Entry
Database: PDB / ID: 8p2v
TitleNeisseria meningitidis Type IV pilus SB-GATDH variant
ComponentsNeisseria meningitidis PilE variant SB-GATDH
KeywordsPROTEIN FIBRIL / Pilin / Extracellular / Adhesion / Aggregation
Function / homologySN-GLYCEROL-3-PHOSPHATE / :
Function and homology information
Biological speciesNeisseria meningitidis 8013 (bacteria)
MethodELECTRON MICROSCOPY / helical reconstruction / cryo EM / Resolution: 2.99 Å
AuthorsFernandez-Martinez, D. / Dumenil, G.
Funding support France, 1items
OrganizationGrant numberCountry
Agence Nationale de la Recherche (ANR)ANR 18 CE11 0022 France
CitationJournal: Nat Commun / Year: 2024
Title: Cryo-EM structures of type IV pili complexed with nanobodies reveal immune escape mechanisms.
Authors: David Fernandez-Martinez / Youxin Kong / Sylvie Goussard / Agustin Zavala / Pauline Gastineau / Martial Rey / Gabriel Ayme / Julia Chamot-Rooke / Pierre Lafaye / Matthijn Vos / Ariel Mechaly ...Authors: David Fernandez-Martinez / Youxin Kong / Sylvie Goussard / Agustin Zavala / Pauline Gastineau / Martial Rey / Gabriel Ayme / Julia Chamot-Rooke / Pierre Lafaye / Matthijn Vos / Ariel Mechaly / Guillaume Duménil /
Abstract: Type IV pili (T4P) are prevalent, polymeric surface structures in pathogenic bacteria, making them ideal targets for effective vaccines. However, bacteria have evolved efficient strategies to evade ...Type IV pili (T4P) are prevalent, polymeric surface structures in pathogenic bacteria, making them ideal targets for effective vaccines. However, bacteria have evolved efficient strategies to evade type IV pili-directed antibody responses. Neisseria meningitidis are prototypical type IV pili-expressing Gram-negative bacteria responsible for life threatening sepsis and meningitis. This species has evolved several genetic strategies to modify the surface of its type IV pili, changing pilin subunit amino acid sequence, nature of glycosylation and phosphoforms, but how these modifications affect antibody binding at the structural level is still unknown. Here, to explore this question, we determine cryo-electron microscopy (cryo-EM) structures of pili of different sequence types with sufficiently high resolution to visualize posttranslational modifications. We then generate nanobodies directed against type IV pili which alter pilus function in vitro and in vivo. Cyro-EM in combination with molecular dynamics simulation of the nanobody-pilus complexes reveals how the different types of pili surface modifications alter nanobody binding. Our findings shed light on the impressive complementarity between the different strategies used by bacteria to avoid antibody binding. Importantly, we also show that structural information can be used to make informed modifications in nanobodies as countermeasures to these immune evasion mechanisms.
History
DepositionMay 16, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Apr 3, 2024Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Neisseria meningitidis PilE variant SB-GATDH
hetero molecules


Theoretical massNumber of molelcules
Total (without water)17,5323
Polymers17,0671
Non-polymers4642
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein Neisseria meningitidis PilE variant SB-GATDH


Mass: 17067.182 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Neisseria meningitidis 8013 (bacteria)
#2: Chemical ChemComp-G3P / SN-GLYCEROL-3-PHOSPHATE / Glycerol 3-phosphate


Mass: 172.074 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C3H9O6P / Feature type: SUBJECT OF INVESTIGATION
#3: Chemical ChemComp-WKE / (2~{R})-~{N}-[(2~{R},3~{S},4~{S},5~{R},6~{R})-5-acetamido-2-methyl-4,6-bis(oxidanyl)oxan-3-yl]-2,3-bis(oxidanyl)propanamide


Mass: 292.286 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C11H20N2O7 / Feature type: SUBJECT OF INVESTIGATION
Has ligand of interestY

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: FILAMENT / 3D reconstruction method: helical reconstruction

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Sample preparation

ComponentName: PilE variant SB containing GATDH and G3P PTMs / Type: COMPLEX / Entity ID: #1 / Source: NATURAL
Molecular weightExperimental value: NO
Source (natural)Organism: Neisseria meningitidis 8013 (bacteria)
Buffer solutionpH: 7.4
SpecimenConc.: 0.01 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 3000 nm / Nominal defocus min: 800 nm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

Software
NameVersionClassificationNB
phenix.real_space_refine1.20.1_4487refinement
PHENIX1.20.1_4487refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Helical symmertyAngular rotation/subunit: 100.7 ° / Axial rise/subunit: 10.433 Å / Axial symmetry: C1
3D reconstructionResolution: 2.99 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 271827 / Symmetry type: HELICAL
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 51.9 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00181243
ELECTRON MICROSCOPYf_angle_d0.40971692
ELECTRON MICROSCOPYf_chiral_restr0.0389205
ELECTRON MICROSCOPYf_plane_restr0.0027213
ELECTRON MICROSCOPYf_dihedral_angle_d3.3569186

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