|Entry||Database: PDB / ID: 5v5s|
|Title||multi-drug efflux; membrane transport; RND superfamily; Drug resistance|
|Keywords||MEMBRANE PROTEIN / multi-drug efflux / membrane transport / RND superfamily / Drug resistance|
|Function/homology||Type I secretion outer membrane protein, TolC / enterobactin transport / Outer membrane efflux protein / RND efflux pump, membrane fusion protein, barrel-sandwich domain / HlyD membrane-fusion protein of T1SS / RND efflux pump, membrane fusion protein / Barrel-sandwich domain of CusB or HlyD membrane-fusion / Outer membrane efflux protein / drug:proton antiporter activity / Hydrophobe/amphiphile efflux-1 HAE1 ...Type I secretion outer membrane protein, TolC / enterobactin transport / Outer membrane efflux protein / RND efflux pump, membrane fusion protein, barrel-sandwich domain / HlyD membrane-fusion protein of T1SS / RND efflux pump, membrane fusion protein / Barrel-sandwich domain of CusB or HlyD membrane-fusion / Outer membrane efflux protein / drug:proton antiporter activity / Hydrophobe/amphiphile efflux-1 HAE1 / efflux pump complex / Acriflavin resistance protein / Multidrug efflux transporter AcrB TolC docking domain, DN/DC subdomains / efflux transmembrane transporter activity / drug transmembrane transport / drug transmembrane transporter activity / AcrB/AcrD/AcrF family / intrinsic component of plasma membrane / porin activity / transporter activity / Prokaryotic membrane lipoprotein lipid attachment site profile. / bile acid and bile salt transport / ion transmembrane transport / cell outer membrane / response to organic cyclic compound / transmembrane transport / outer membrane-bounded periplasmic space / protein N-terminus binding / ion channel activity / response to drug / response to antibiotic / integral component of plasma membrane / membrane / integral component of membrane / identical protein binding / plasma membrane / Outer membrane protein TolC / Multidrug efflux pump subunit AcrA / Multidrug efflux pump subunit AcrA / Multidrug efflux pump subunit AcrB|
Function and homology information
|Specimen source||Escherichia coli / / bacteria /|
|Method||Electron microscopy (6.5 Å resolution / Particle / Single particle) / Transmission electron microscopy|
|Authors||wang, Z. / fan, G. / Hryc, C.F. / Blaza, J.N. / Serysheva, I.I. / Schmid, M.F. / Chiu, W. / Luisi, B.F. / Du, D.|
|Citation||Journal: 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.
SummaryFull reportAbout validation report
|Date||Deposition: Mar 15, 2017 / Release: Apr 19, 2017|
Downloads & links
A: Outer membrane protein TolC
B: Outer membrane protein TolC
C: Outer membrane protein TolC
D: Multidrug efflux pump subunit AcrA
E: Multidrug efflux pump subunit AcrA
F: Multidrug efflux pump subunit AcrA
G: Multidrug efflux pump subunit AcrA
H: Multidrug efflux pump subunit AcrA
I: Multidrug efflux pump subunit AcrA
J: Multidrug efflux pump subunit AcrB
K: Multidrug efflux pump subunit AcrB
L: Multidrug efflux pump subunit AcrB
Mass: 48673.801 Da / Num. of mol.: 3 / Source: (gene. exp.) Escherichia coli / / bacteria /
Gene: tolC, colE1-i, mtcB, mukA, refI, toc, weeA, b3035, JW5503
Production host: Escherichia coli / References: UniProt:P02930
Mass: 42253.551 Da / Num. of mol.: 6 / Source: (gene. exp.) Escherichia coli / / bacteria / / Gene: acrA, Z0578, ECs0516 / Production host: Escherichia coli / References: UniProt:P0AE07, UniProt:P0AE06*PLUS
Mass: 113681.242 Da / Num. of mol.: 3 / Source: (gene. exp.) Escherichia coli / / bacteria / / Gene: acrB, acrE, b0462, JW0451 / Production host: Escherichia coli / References: UniProt:P31224
|Experiment||Method: ELECTRON MICROSCOPY|
|EM experiment||Aggregation state: PARTICLE / Reconstruction method: SINGLE PARTICLE|
|Component||Name: AcrABTolC in apo state / Type: COMPLEX / Entity ID: 1,||Source (natural)||Organism: Escherichia coli||Source (recombinant)||Organism: Escherichia coli||Buffer solution||pH: 8||Specimen||Conc.: 2 mg/ml / Details: AcrABTolC complex in apo state / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES||Specimen support||Grid material: GOLD / Grid mesh size: 300 / Grid type: Quantifoil||Vitrification||Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 20 kelvins|
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)
-Electron microscopy imaging
Model: Titan Krios / Image courtesy: FEI Company
|Microscopy||Microscope model: FEI TITAN KRIOS|
|Electron gun||Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM|
|Electron lens||Mode: BRIGHT FIELD|
|Image recording||Electron dose: 1 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k)|
|Image scans||Movie frames/image: 45|
|Software||Name: PHENIX / Version: dev_2415: / Classification: refinement|
|CTF correction||Type: PHASE FLIPPING ONLY|
|Particle selection||Details: auto box by relion / Number of particles selected: 95410|
|Symmetry||Point symmetry: C3|
|3D reconstruction||Resolution: 6.5 Å / Resolution method: FSC 0.143 CUT-OFF / Number of particles: 13544 / Algorithm: BACK PROJECTION / Number of class averages: 1 / Symmetry type: POINT|
|Atomic model building||Details: Crystal 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.|
Ref protocol: FLEXIBLE FIT / Ref space: REAL / Target criteria: Cross-correlation coefficient
|Refine LS restraints|
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