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- EMDB-10371: CryoEM structure of MexB in nanodisc -

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Basic information

Entry
Database: EMDB / ID: EMD-10371
TitleCryoEM structure of MexB in nanodisc
Map dataCryoEM structure of RND transport MexB trimer from Pseudomonas aeruginosa stabilized in nanodisc
Sample
  • Complex: MexB
    • Protein or peptide: MEXB
Function / homology
Function and homology information


efflux transmembrane transporter activity / xenobiotic transmembrane transporter activity / transmembrane transport / response to antibiotic / plasma membrane
Similarity search - Function
Hydrophobe/amphiphile efflux-1 HAE1 / Acriflavin resistance protein / Multidrug efflux transporter AcrB TolC docking domain, DN/DC subdomains / AcrB/AcrD/AcrF family
Similarity search - Domain/homology
Efflux pump membrane transporter / Multidrug resistance protein MexB
Similarity search - Component
Biological speciesPseudomonas aeruginosa (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.5 Å
AuthorsGlavier M / Puvanendran D / Salvador D / Decossas M / Phan G / Garnier C / Frezza E / Cece Q / Schoehn G / Picard M ...Glavier M / Puvanendran D / Salvador D / Decossas M / Phan G / Garnier C / Frezza E / Cece Q / Schoehn G / Picard M / Taveau J-C / Daury L / Broutin I / Lambert O
Funding support France, 1 items
OrganizationGrant numberCountry
French National Research AgencyANR-17CE11-0028 France
CitationJournal: Nat Commun / Year: 2020
Title: Antibiotic export by MexB multidrug efflux transporter is allosterically controlled by a MexA-OprM chaperone-like complex.
Authors: Marie Glavier / Dhenesh Puvanendran / Dimitri Salvador / Marion Decossas / Gilles Phan / Cyril Garnier / Elisa Frezza / Quentin Cece / Guy Schoehn / Martin Picard / Jean-Christophe Taveau / ...Authors: Marie Glavier / Dhenesh Puvanendran / Dimitri Salvador / Marion Decossas / Gilles Phan / Cyril Garnier / Elisa Frezza / Quentin Cece / Guy Schoehn / Martin Picard / Jean-Christophe Taveau / Laetitia Daury / Isabelle Broutin / Olivier Lambert /
Abstract: The tripartite multidrug efflux system MexAB-OprM is a major actor in Pseudomonas aeruginosa antibiotic resistance by exporting a large variety of antimicrobial compounds. Crystal structures of MexB ...The tripartite multidrug efflux system MexAB-OprM is a major actor in Pseudomonas aeruginosa antibiotic resistance by exporting a large variety of antimicrobial compounds. Crystal structures of MexB and of its Escherichia coli homolog AcrB had revealed asymmetric trimers depicting a directional drug pathway by a conformational interconversion (from Loose and Tight binding pockets to Open gate (LTO) for drug exit). It remains unclear how MexB acquires its LTO form. Here by performing functional and cryo-EM structural investigations of MexB at various stages of the assembly process, we unveil that MexB inserted in lipid membrane is not set for active transport because it displays an inactive LTC form with a Closed exit gate. In the tripartite complex, OprM and MexA form a corset-like platform that converts MexB into the active form. Our findings shed new light on the resistance nodulation cell division (RND) cognate partners which act as allosteric factors eliciting the functional drug extrusion.
History
DepositionOct 9, 2019-
Header (metadata) releaseOct 14, 2020-
Map releaseOct 14, 2020-
UpdateApr 14, 2021-
Current statusApr 14, 2021Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.02
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.02
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6t7s
  • Surface level: 0.02
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_10371.png

Downloads & links

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Map

FileDownload / File: emd_10371.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationCryoEM structure of RND transport MexB trimer from Pseudomonas aeruginosa stabilized in nanodisc
Voxel sizeX=Y=Z: 1.36 Å
Density
Contour LevelBy AUTHOR: 0.02 / Movie #1: 0.02
Minimum - Maximum-0.029316468 - 0.067547865
Average (Standard dev.)-3.3355784e-06 (±0.0027080537)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 408.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.361.361.36
M x/y/z300300300
origin x/y/z0.0000.0000.000
length x/y/z408.000408.000408.000
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS300300300
D min/max/mean-0.0290.068-0.000

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Supplemental data

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Sample components

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Entire : MexB

EntireName: MexB
Components
  • Complex: MexB
    • Protein or peptide: MEXB

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Supramolecule #1: MexB

SupramoleculeName: MexB / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Pseudomonas aeruginosa (bacteria)
Recombinant expressionOrganism: Escherichia coli (E. coli)

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Macromolecule #1: MEXB

MacromoleculeName: MEXB / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Pseudomonas aeruginosa (bacteria)
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MSKFFIDRPI FAWVIALVIM LAGGLSILSL PVNQYPAIAP PAIAVQVSYP GASAETVQDT VVQVIEQQM NGIDNLRYIS SESNSDGSMT ITVTFEQGTD PDIAQVQVQN KLQLATPLLP Q EVQRQGIR VTKAVKNFLM VVGVVSTDGS MTKEDLSNYI VSNIQDPLSR ...String:
MSKFFIDRPI FAWVIALVIM LAGGLSILSL PVNQYPAIAP PAIAVQVSYP GASAETVQDT VVQVIEQQM NGIDNLRYIS SESNSDGSMT ITVTFEQGTD PDIAQVQVQN KLQLATPLLP Q EVQRQGIR VTKAVKNFLM VVGVVSTDGS MTKEDLSNYI VSNIQDPLSR TKGVGDFQVF GS QYSMRIW LDPAKLNSYQ LTPGDVSSAI QAQNVQISSG QLGGLPAVKG QQLNATIIGK TRL QTAEQF ENILLKVNPD GSQVRLKDVA DVGLGGQDYS INAQFNGSPA SGIAIKLATG ANAL DTAKA IRQTIANLEP FMPQGMKVVY PYDTTPVVSA SIHEVVKTLG EAILLVFLVM YLFLQ NFRA TLIPTIAVPV VLLGTFGVLA AFGFSINTLT MFGMVLAIGL LVDDAIVVVE NVERVM AEE GLSPREAARK SMGQIQGALV GIAMVLSAVF LPMAFFGGST GVIYRQFSIT IVSAMAL SV IVALILTPAL CATMLKPIEK GDHGEHKGGF FGWFNRMFLS TTHGYERGVA SILKHRAP Y LLIYVVIVAG MIWMFTRIPT AFLPDEDQGV LFAQVQTPPG SSAERTQVVV DSMREYLLE KESSSVSSVF TVTGFNFAGR GQSSGMAFIM LKPWEERPGG ENSVFELAKR AQMHFFSFKD AMVFAFAPP SVLELGNATG FDLFLQDQAG VGHEVLLQAR NKFLMLAAQN PALQRVRPNG M SDEPQYKL EIDDEKASAL GVSLADINST VSIAWGSSYV NDFIDRGRVK RVYLQGRPDA RM NPDDLSK WYVRNDKGEM VPFNAFATGK WEYGSPKLER YNGVPAMEIL GEPAPGLSSG DAM AAVEEI VKQLPKGVGY SWTGLSYEER LSGSQAPALY ALSLLVVFLC LAALYESWSI PFSV MLVVP LGVIGALLAT SMRGLSNDVF FQVGLLTTIG LSAKNAILIV EFAKELHEQG KGIVE AAIE ACRMRLRPIV MTSLAFILGV VPLAISTGAG SGSQHAIGTG VIGGMVTATV LAIFWV PLF YVAVSTLFKD EASKQQASVE KGQ

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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: 7.4
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 QUANTUM (4k x 4k) / Average electron dose: 42.0 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: 212667
CTF correctionSoftware - Name: Gctf
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 4.5 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION / Number images used: 41604
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION
FSC plot (resolution estimation)

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