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- PDB-7kgd: Cryo-EM Structures of AdeB from Acinetobacter baumannii: AdeB-I -

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Basic information

Entry
Database: PDB / ID: 7kgd
TitleCryo-EM Structures of AdeB from Acinetobacter baumannii: AdeB-I
ComponentsEfflux pump membrane transporter
KeywordsTRANSPORT PROTEIN / AdeB / Acinetobacter baumannii / Membrane Protein / Cryo-EM / RND transporter
Function / homologyHydrophobe/amphiphile efflux-1 HAE1 / Multidrug efflux transporter AcrB TolC docking domain, DN/DC subdomains / Acriflavin resistance protein / AcrB/AcrD/AcrF family / efflux transmembrane transporter activity / xenobiotic transmembrane transporter activity / plasma membrane / PHOSPHATIDYLETHANOLAMINE / Efflux pump membrane transporter
Function and homology information
Biological speciesAcinetobacter baumannii (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.64 Å
AuthorsMorgan, C.E. / Yu, E.W.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID) United States
CitationJournal: mBio / Year: 2021
Title: Cryoelectron Microscopy Structures of AdeB Illuminate Mechanisms of Simultaneous Binding and Exporting of Substrates.
Authors: Christopher E Morgan / Przemyslaw Glaza / Inga V Leus / Anhthu Trinh / Chih-Chia Su / Meng Cui / Helen I Zgurskaya / Edward W Yu /
Abstract: is a Gram-negative pathogen that has emerged as one of the most highly antibiotic-resistant bacteria worldwide. Multidrug efflux within these highly drug-resistant strains and other opportunistic ... is a Gram-negative pathogen that has emerged as one of the most highly antibiotic-resistant bacteria worldwide. Multidrug efflux within these highly drug-resistant strains and other opportunistic pathogens is a major cause of failure of drug-based treatments of infectious diseases. The best-characterized multidrug efflux system in is the prevalent rug fflux B (AdeB) pump, which is a member of the resistance-nodulation-cell division (RND) superfamily. Here, we report six structures of the trimeric AdeB multidrug efflux pump in the presence of ethidium bromide using single-particle cryoelectron microscopy (cryo-EM). These structures allow us to directly observe various novel conformational states of the AdeB trimer, including the transmembrane region of trimeric AdeB can be associated with form a trimer assembly or dissociated into "dimer plus monomer" and "monomer plus monomer plus monomer" configurations. We also discover that a single AdeB protomer can simultaneously anchor a number of ethidium ligands and that different AdeB protomers can bind ethidium molecules simultaneously. Combined with molecular dynamics (MD) simulations, we reveal a drug transport mechanism that involves multiple multidrug-binding sites and various transient states of the AdeB membrane protein. Our data suggest that each AdeB protomer within the trimer binds and exports drugs independently. has emerged as one of the most highly antibiotic-resistant Gram-negative pathogens. The prevalent AdeB multidrug efflux pump mediates resistance to a broad spectrum of clinically relevant antimicrobial agents. Here, we report six cryo-EM structures of the trimeric AdeB pump in the presence of ethidium bromide. We discover that a single AdeB protomer can simultaneously anchor a number of ligands, and different AdeB protomers can bind ethidium molecules simultaneously. The results indicate that each AdeB protomer within the trimer recognizes and extrudes drugs independently.
History
DepositionOct 16, 2020Deposition site: RCSB / Processing site: RCSB
Revision 1.0Mar 10, 2021Provider: repository / Type: Initial release
Revision 1.1Mar 6, 2024Group: Data collection / Database references / Category: chem_comp_atom / chem_comp_bond / database_2
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession

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Assembly

Deposited unit
A: Efflux pump membrane transporter
B: Efflux pump membrane transporter
C: Efflux pump membrane transporter
hetero molecules


Theoretical massNumber of molelcules
Total (without water)342,1699
Polymers337,7653
Non-polymers4,4046
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: microscopy
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area20670 Å2
ΔGint-126 kcal/mol
Surface area121740 Å2

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Components

#1: Protein Efflux pump membrane transporter


Mass: 112588.297 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Acinetobacter baumannii (bacteria) / Gene: adeB, acrB, 32_436, CBI29_01998, EA686_00900 / Production host: Escherichia coli (E. coli) / References: UniProt: Q2FD70
#2: Chemical
ChemComp-PTY / PHOSPHATIDYLETHANOLAMINE


Mass: 734.039 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: C40H80NO8P / Comment: phospholipid*YM
Has ligand of interestN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: AdeB / Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Source (natural)Organism: Acinetobacter baumannii (bacteria)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 6.5
SpecimenEmbedding 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: SPOT SCAN
Electron lensMode: BRIGHT FIELD
Image recordingElectron dose: 40 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K2 SUMMIT (4k x 4k)

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Processing

EM softwareName: cryoSPARC / Version: 2.15 / Category: 3D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C3 (3 fold cyclic)
3D reconstructionResolution: 3.64 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 16223 / Symmetry type: POINT

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