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- EMDB-8636: multi-drug efflux; membrane transport; RND superfamily; Drug resi... -

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Basic information

Entry
Database: EMDB / ID: EMD-8636
Titlemulti-drug efflux; membrane transport; RND superfamily; Drug resistance
Map dataMutant cryoEM density map.
Sample
  • Complex: AcrABTolC in apo state
    • Protein or peptide: Outer membrane protein TolC
    • Protein or peptide: Multidrug efflux pump subunit AcrA
    • Protein or peptide: Multidrug efflux pump subunit AcrB
Keywordsmulti-drug efflux / membrane transport / RND superfamily / Drug resistance / MEMBRANE PROTEIN
Function / homology
Function and homology information


MacAB-TolC complex / alkane transmembrane transporter activity / alkane transport / xenobiotic detoxification by transmembrane export across the cell outer membrane / efflux pump complex / enterobactin transport / enterobactin transmembrane transporter activity / periplasmic side of plasma membrane / xenobiotic detoxification by transmembrane export across the plasma membrane / bile acid transmembrane transporter activity ...MacAB-TolC complex / alkane transmembrane transporter activity / alkane transport / xenobiotic detoxification by transmembrane export across the cell outer membrane / efflux pump complex / enterobactin transport / enterobactin transmembrane transporter activity / periplasmic side of plasma membrane / xenobiotic detoxification by transmembrane export across the plasma membrane / bile acid transmembrane transporter activity / bile acid and bile salt transport / xenobiotic transport / porin activity / monoatomic ion channel activity / efflux transmembrane transporter activity / xenobiotic transmembrane transporter activity / transmembrane transporter activity / fatty acid transport / cell outer membrane / : / response to toxic substance / outer membrane-bounded periplasmic space / monoatomic ion transmembrane transport / response to xenobiotic stimulus / response to antibiotic / identical protein binding / membrane / plasma membrane
Similarity search - Function
: / Type I secretion outer membrane protein, TolC / : / Cation efflux system protein CusB, domain 1 / Cation efflux system protein CusB domain 1 / RND efflux pump, membrane fusion protein / RND efflux pump, membrane fusion protein, barrel-sandwich domain / Barrel-sandwich domain of CusB or HlyD membrane-fusion / Outer membrane efflux protein / Outer membrane efflux protein ...: / Type I secretion outer membrane protein, TolC / : / Cation efflux system protein CusB, domain 1 / Cation efflux system protein CusB domain 1 / RND efflux pump, membrane fusion protein / RND efflux pump, membrane fusion protein, barrel-sandwich domain / Barrel-sandwich domain of CusB or HlyD membrane-fusion / Outer membrane efflux protein / Outer membrane efflux protein / Hydrophobe/amphiphile efflux-1 HAE1 / Acriflavin resistance protein / Multidrug efflux transporter AcrB TolC docking domain, DN/DC subdomains / AcrB/AcrD/AcrF family / Prokaryotic membrane lipoprotein lipid attachment site profile.
Similarity search - Domain/homology
Outer membrane protein TolC / Multidrug efflux pump subunit AcrA / Multidrug efflux pump subunit AcrA / Multidrug efflux pump subunit AcrB
Similarity search - Component
Biological speciesEscherichia coli (E. coli) / Escherichia coli
Methodsingle particle reconstruction / cryo EM / Resolution: 6.5 Å
Authorswang Z / fan G
CitationJournal: Elife / Year: 2017
Title: An allosteric transport mechanism for the AcrAB-TolC multidrug efflux pump.
Authors: Zhao Wang / Guizhen Fan / Corey F Hryc / James N Blaza / Irina I Serysheva / Michael F Schmid / Wah Chiu / Ben F Luisi / Dijun Du /
Abstract: Bacterial efflux pumps confer multidrug resistance by transporting diverse antibiotics from the cell. In Gram-negative bacteria, some of these pumps form multi-protein assemblies that span the cell ...Bacterial efflux pumps confer multidrug resistance by transporting diverse antibiotics from the cell. In Gram-negative bacteria, some of these pumps form multi-protein assemblies that span the cell envelope. Here, we report the near-atomic resolution cryoEM structures of the AcrAB-TolC multidrug efflux pump in resting and drug transport states, revealing a quaternary structural switch that allosterically couples and synchronizes initial ligand binding with channel opening. Within the transport-activated state, the channel remains open even though the pump cycles through three distinct conformations. Collectively, our data provide a dynamic mechanism for the assembly and operation of the AcrAB-TolC pump.
History
DepositionMar 15, 2017-
Header (metadata) releaseApr 19, 2017-
Map releaseApr 19, 2017-
UpdateNov 20, 2024-
Current statusNov 20, 2024Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0266
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.0266
  • Imaged by UCSF Chimera
  • Download
  • Surface view with fitted model
  • Atomic models: PDB-5v5s
  • Surface level: 0.0266
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_8636.png

Downloads & links

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Map

FileDownload / File: emd_8636.map.gz / Format: CCP4 / Size: 209.3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationMutant cryoEM density map.
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.06 Å/pix.
x 380 pix.
= 402.8 Å		1.06 Å/pix.
x 380 pix.
= 402.8 Å		1.06 Å/pix.
x 380 pix.
= 402.8 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.06 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0266 / Movie #1: 0.0266
Minimum - Maximum-0.030838259 - 0.09197038
Average (Standard dev.)0.00056815404 (±0.0040272484)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions380380380
Spacing380380380
CellA=B=C: 402.8 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.061.061.06
M x/y/z380380380
origin x/y/z0.0000.0000.000
length x/y/z402.800402.800402.800
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS380380380
D min/max/mean-0.0310.0920.001

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Supplemental data

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Sample components

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Entire : AcrABTolC in apo state

EntireName: AcrABTolC in apo state
Components
  • Complex: AcrABTolC in apo state
    • Protein or peptide: Outer membrane protein TolC
    • Protein or peptide: Multidrug efflux pump subunit AcrA
    • Protein or peptide: Multidrug efflux pump subunit AcrB

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Supramolecule #1: AcrABTolC in apo state

SupramoleculeName: AcrABTolC in apo state / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Escherichia coli (E. coli)

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Macromolecule #1: Outer membrane protein TolC

MacromoleculeName: Outer membrane protein TolC / type: protein_or_peptide / ID: 1 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 48.673801 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: ENL(MSE)QVYQQA RLSNPELRKS AADRDAAFEK INEARSPLLP QLGLGADYTY SNGYRDANGI NSNATSASLQ LTQSIF D(MSE)S KWRALTLQEK AAGIQDVTYQ TDQQTLILNT ATAYFNVLNA IDVLSYTQAQ KEAIYRQLDQ TTQRFNVGLV AIT DVQNAR ...String:
ENL(MSE)QVYQQA RLSNPELRKS AADRDAAFEK INEARSPLLP QLGLGADYTY SNGYRDANGI NSNATSASLQ LTQSIF D(MSE)S KWRALTLQEK AAGIQDVTYQ TDQQTLILNT ATAYFNVLNA IDVLSYTQAQ KEAIYRQLDQ TTQRFNVGLV AIT DVQNAR AQYDTVLANE VTARNNLDNA VEQLRQITGN YYPELAALNV ENFKTDKPQP VNALLKEAEK RNLSLLQARL SQDL AREQI RQAQDGHLPT LDLTASTGIS DTSYSGSKTR GAAGTQYDDS N(MSE)GQNKVGLS FSLPIYQGG(MSE) VNSQVKQ AQ YNFVGASEQL ESAHRSVVQT VRSSFNNINA SISSINAYKQ AVVSAQSSLD A(MSE)EAGYSVGT RTIVDVLDAT TTLY NAKQE LANARYNYLI NQLNIKSALG TLNEQDLLAL NNALSKPVST NPENVAPQTP EQNAIAD

UniProtKB: Outer membrane protein TolC

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Macromolecule #2: Multidrug efflux pump subunit AcrA

MacromoleculeName: Multidrug efflux pump subunit AcrA / type: protein_or_peptide / ID: 2 / Number of copies: 6 / Enantiomer: LEVO
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 42.253551 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MNKNRGFTPL AVVLMLSGSL ALTGCDDKQA QQGGQQMPAV GVVTVKTEPL QITTELPGRT SAYRIAEVRP QVSGIILKRN FKEGSDIEA GVSLYQIDPA TYQATYDSAK GDLAKAQAAA NIAQLTVNRY QKLLGTQYIS KQEYDQALAD AQQANAAVTA A KAAVETAR ...String:
MNKNRGFTPL AVVLMLSGSL ALTGCDDKQA QQGGQQMPAV GVVTVKTEPL QITTELPGRT SAYRIAEVRP QVSGIILKRN FKEGSDIEA GVSLYQIDPA TYQATYDSAK GDLAKAQAAA NIAQLTVNRY QKLLGTQYIS KQEYDQALAD AQQANAAVTA A KAAVETAR INLAYTKVTS PISGRIGKSN VTEGALVQNG QATALATVQQ LDPIYVDVTQ SSNDFLRLKQ ELANGTLKQE NG KAKVSLI TSDGIKFPQD GTLEFSDVTV DQTTGCITLR AIFPNPDHTL LPGMFVRARL EEGLNPNAIL VPQQGVTRTP RGD ATVLVV GADDKVETRP IVASQAIGDK WLVTEGLKAG DRVVISGLQK VRPGVQVKAQ EVTADNNQQA ASGAQPEQSK S

UniProtKB: Multidrug efflux pump subunit AcrA

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Macromolecule #3: Multidrug efflux pump subunit AcrB

MacromoleculeName: Multidrug efflux pump subunit AcrB / type: protein_or_peptide / ID: 3 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Escherichia coli
Molecular weightTheoretical: 113.681242 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MPNFFIDRPI FAWVIAIIIM LAGGLAILKL PVAQYPTIAP PAVTISASYP GADAKTVQDT VTQVIEQNMN GIDNLMYMSS NSDSTGTVQ ITLTFESGTD ADIAQVQVQN KLQLAMPLLP QEVQQQGVSV EKSSSSFLMV VGVINTDGTM TQEDISDYVA A NMKDAISR ...String:
MPNFFIDRPI FAWVIAIIIM LAGGLAILKL PVAQYPTIAP PAVTISASYP GADAKTVQDT VTQVIEQNMN GIDNLMYMSS NSDSTGTVQ ITLTFESGTD ADIAQVQVQN KLQLAMPLLP QEVQQQGVSV EKSSSSFLMV VGVINTDGTM TQEDISDYVA A NMKDAISR TSGVGDVQLF GSQYAMRIWM NPNELNKFQL TPVDVITAIK AQNAQVAAGQ LGGTPPVKGQ QLNASIIAQT RL TSTEEFG KILLKVNQDG CRVLLRDVAK IELGGENYDI IAEFNGQPAS GLGIKLATGA NALDTAAAIR AELAKMEPFF PSG LKIVYP YDTTPFVKIS IHEVVKTLVE AIILVFLVMY LFLQNFRATL IPTIAVPVVL LGTFAVLAAF GFSINTLTMF GMVL AIGLL VDDAIVVVEN VERVMAEEGL PPKEATRKSM GQIQGALVGI AMVLSAVFVP MAFFGGSTGA IYRQFSITIV SAMAL SVLV ALILTPALCA TMLKPIAKGD HGEGKKGFFG WFNRMFEKST HHYTDSVGGI LRSTGRYLVL YLIIVVGMAY LFVRLP SSF LPDEDQGVFM TMVQLPAGAT QERTQKVLNE VTHYYLTKEK NNVESVFAVN GFGFAGRGQN TGIAFVSLKD WADRPGE EN KVEAITMRAT RAFSQIKDAM VFAFNLPAIV ELGTATGFDF ELIDQAGLGH EKLTQARNQL LAEAAKHPDM LTSVRPNG L EDTPQFKIDI DQEKAQALGV SINDINTTLG AAWGGSYVND FIDRGRVKKV YVMSEAKYRM LPDDIGDWYV RAADGQMVP FSAFSSSRWE YGSPRLERYN GLPSMEILGQ AAPGKSTGEA MELMEQLASK LPTGVGYDWT GMSYQERLSG NQAPSLYAIS LIVVFLCLA ALYESWSIPF SVMLVVPLGV IGALLAATFR GLTNDVYFQV GLLTTIGLSA KNAILIVEFA KDLMDKEGKG L IEATLDAV RMRLRPILMT SLAFILGVMP LVISTGAGSG AQNAVGTGVM GGMVTATVLA IFFVPVFFVV VRRRFSRKNE DI EHSHTVD HH

UniProtKB: Multidrug efflux pump subunit AcrB

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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

Concentration2 mg/mL
BufferpH: 8
GridModel: Quantifoil / Material: GOLD / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 20 K / Instrument: FEI VITROBOT MARK IV
Details: a 3ul aliquot at a concentration of 2 mg per ml was applied onto glow-discharged holey carbon grid (Quantifoil Au R1.21.3, 300 mesh).
DetailsAcrABTolC complex in apo state

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Average electron dose: 1.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 95410 / Details: auto box by relion
Startup modelType of model: NONE / Details: generate from scratch by EMAN2
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C3 (3 fold cyclic) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 6.5 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 13544
Initial angle assignmentType: COMMON LINE / Software - Name: EMAN (ver. 2.2)
Details: e2initialmodel.py generate from reference free 2d averages
Final angle assignmentType: PROJECTION MATCHING / Software - Name: RELION (ver. 2)
Final 3D classificationNumber classes: 1
FSC plot (resolution estimation)

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Atomic model buiding 1

DetailsCrystal structures were rigid-body fit into the density map and model optimization was then carried out with Phenix real-space refine. Due to the weaker resolution, stronger stereochemical and secondary structure restraints were used to ensure that alpha-helices and beta-sheets did not deviate far from their expected geometry. Manual adjustments were kept to a minimum to reduce human bias in the modeling procedure, with Coot only being used to fix obvious errors such as C-beta deviations. A final check of MolProbity and cross correlation was done to ensure model quality.
RefinementSpace: REAL / Protocol: FLEXIBLE FIT / Target criteria: Cross-correlation coefficient
Output model

PDB-5v5s:
multi-drug efflux; membrane transport; RND superfamily; Drug resistance

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