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- EMDB-12089: Acinetobacter baumannii multidrug transporter AdeB in L*OO state -

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Basic information

Entry
Database: EMDB / ID: EMD-12089
TitleAcinetobacter baumannii multidrug transporter AdeB in L*OO state
Map data
Sample
  • Complex: Homotrimeric RND-transporter AdeB reconstituted in Salipro
    • Protein or peptide: Efflux pump membrane transporter
KeywordsRND-transporter / multidrug transporter / antibiotic resistance / membrane protein / TRANSPORT PROTEIN
Function / homologyHydrophobe/amphiphile efflux-1 HAE1 / Multidrug efflux transporter AcrB TolC docking domain, DN/DC subdomains / Acriflavin resistance protein / AcrB/AcrD/AcrF family / xenobiotic transport / efflux transmembrane transporter activity / response to toxic substance / plasma membrane / Efflux pump membrane transporter
Function and homology information
Biological speciesAcinetobacter baumannii AYE (bacteria) / Acinetobacter baumannii (strain AYE) (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.84 Å
AuthorsOrnik-Cha A / Reitz J
Funding support Germany, European Union, 5 items
OrganizationGrant numberCountry
German Research Foundation (DFG)SFB807 Germany
German Research Foundation (DFG)EXC115 Germany
German Research Foundation (DFG)FR-1653/12 Germany
German Research Foundation (DFG)SFB902 Germany
Innovative Medicines Initiative115525European Union
CitationJournal: Nat Commun / Year: 2021
Title: Structural and functional analysis of the promiscuous AcrB and AdeB efflux pumps suggests different drug binding mechanisms.
Authors: Alina Ornik-Cha / Julia Wilhelm / Jessica Kobylka / Hanno Sjuts / Attilio V Vargiu / Giuliano Malloci / Julian Reitz / Anja Seybert / Achilleas S Frangakis / Klaas M Pos /
Abstract: Upon antibiotic stress Gram-negative pathogens deploy resistance-nodulation-cell division-type tripartite efflux pumps. These include a H/drug antiporter module that recognizes structurally diverse ...Upon antibiotic stress Gram-negative pathogens deploy resistance-nodulation-cell division-type tripartite efflux pumps. These include a H/drug antiporter module that recognizes structurally diverse substances, including antibiotics. Here, we show the 3.5 Å structure of subunit AdeB from the Acinetobacter baumannii AdeABC efflux pump solved by single-particle cryo-electron microscopy. The AdeB trimer adopts mainly a resting state with all protomers in a conformation devoid of transport channels or antibiotic binding sites. However, 10% of the protomers adopt a state where three transport channels lead to the closed substrate (deep) binding pocket. A comparison between drug binding of AdeB and Escherichia coli AcrB is made via activity analysis of 20 AdeB variants, selected on basis of side chain interactions with antibiotics observed in the AcrB periplasmic domain X-ray co-structures with fusidic acid (2.3 Å), doxycycline (2.1 Å) and levofloxacin (2.7 Å). AdeABC, compared to AcrAB-TolC, confers higher resistance to E. coli towards polyaromatic compounds and lower resistance towards antibiotic compounds.
History
DepositionDec 13, 2020-
Header (metadata) releaseOct 20, 2021-
Map releaseOct 20, 2021-
UpdateJul 10, 2024-
Current statusJul 10, 2024Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.025
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.025
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7b8q
  • Surface level: 0.025
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-7b8q
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_12089.png

Downloads & links

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Map

FileDownload / File: emd_12089.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.05 Å/pix.
x 256 pix.
= 268.8 Å		1.05 Å/pix.
x 256 pix.
= 268.8 Å		1.05 Å/pix.
x 256 pix.
= 268.8 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.05 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.011 / Movie #1: 0.025
Minimum - Maximum-0.07339091 - 0.11627398
Average (Standard dev.)0.00038290164 (±0.0041334922)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 268.8 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.051.051.05
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z268.800268.800268.800
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS256256256
D min/max/mean-0.0730.1160.000

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Supplemental data

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Sample components

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Entire : Homotrimeric RND-transporter AdeB reconstituted in Salipro

EntireName: Homotrimeric RND-transporter AdeB reconstituted in Salipro
Components
  • Complex: Homotrimeric RND-transporter AdeB reconstituted in Salipro
    • Protein or peptide: Efflux pump membrane transporter

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Supramolecule #1: Homotrimeric RND-transporter AdeB reconstituted in Salipro

SupramoleculeName: Homotrimeric RND-transporter AdeB reconstituted in Salipro
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Acinetobacter baumannii AYE (bacteria)
Molecular weightTheoretical: 345 KDa

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Macromolecule #1: Efflux pump membrane transporter

MacromoleculeName: Efflux pump membrane transporter / type: protein_or_peptide / ID: 1 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Acinetobacter baumannii (strain AYE) (bacteria) / Strain: AYE
Molecular weightTheoretical: 115.143055 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MSMSQFFIRR PVFAWVIAIF IIIFGLLSIP KLPIARFPSV APPQVNISAT YPGATAKTIN DSVVTLIERE LSGVKNLLYY SATTDTSGT AEITATFKPG TDVEMAQVDV QNKIKAVEAR LPQVVRQQGL QVEASSSGFL MLVGINSPNN QYSEVDLSDY L VRNVVEEL ...String:
MSMSQFFIRR PVFAWVIAIF IIIFGLLSIP KLPIARFPSV APPQVNISAT YPGATAKTIN DSVVTLIERE LSGVKNLLYY SATTDTSGT AEITATFKPG TDVEMAQVDV QNKIKAVEAR LPQVVRQQGL QVEASSSGFL MLVGINSPNN QYSEVDLSDY L VRNVVEEL KRVEGVGKVQ SFGAEKAMRI WVDPNKLVSY GLSISDVNNA IRENNVEIAP GRLGDLPAEK GQLITIPLSA QG QLSSLEQ FKNISLKSKT NGSVIKLSDV ANVEIGSQAY NFAILENGKP ATAAAIQLSP GANAVKTAEG VRAKIEELKL NLP EGMEFS IPYDTAPFVK ISIEKVIHTL LEAMVLVFIV MYLFLHNVRY TLIPAIVAPI ALLGTFTVML LAGFSINVLT MFGM VLAIG IIVDDAIVVV ENVERIMATE GLSPKDATSK AMKEITSPII GITLVLAAVF LPMAFASGSV GVIYKQFTLT MSVSI LFSA LLALILTPAL CATILKPIDG HHQKKGFFAW FDRSFDKVTK KYELMLLKII KHTVPMMVIF LVITGITFAG MKYWPT AFM PEEDQGWFMT SFQLPSDATA ERTRNVVNQF ENNLKDNPDV KSNTAILGWG FSGAGQNVAV AFTTLKDFKE RTSSASK MT SDVNSSMANS TEGETMAVLP PAIDELGTFS GFSLRLQDRA NLGMPALLAA QDELMAMAAK NKKFYMVWNE GLPQGDNI S LKIDREKLSA LGVKFSDVSD IISTSMGSMY INDFPNQGRM QQVIVQVEAK SRMQLKDILN LKVMGSSGQL VSLSEVVTP QWNKAPQQYN RYNGRPSLSI AGIPNFDTSS GEAMREMEQL IAKLPKGIGY EWTGISLQEK QSESQMAFLL GLSMLVVFLV LAALYESWA IPLSVMLVVP LGIFGAIIAI MSRGLMNDVF FKIGLITIIG LSAKNAILIV EFAKMLKEEG MSLIEATVAA A KLRLRPIL MTSLAFTCGV IPLVIATGAS SETQHALGTG VFGGMISATI LAIFFVPVFF IFILGAVEKL FSSKKKISSA LE VLFQGPH HHHHHHHHH

UniProtKB: Efflux pump membrane transporter

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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

BufferpH: 8.5
Component:
ConcentrationNameFormula
20.0 mMTris-HCl pH 8.5
150.0 mMsodium chlorideNaCl
GridModel: Quantifoil R1.2/1.3 / Material: COPPER
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 60.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

2gif

Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.84 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 34890
Initial angle assignmentType: RANDOM ASSIGNMENT / Software - Name: RELION (ver. 3.0)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0)
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementSpace: REAL
Output model

PDB-7b8q:
Acinetobacter baumannii multidrug transporter AdeB in L*OO state

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