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- EMDB-22844: Cryo-electron microscopy structure of the heavy metal efflux pump... -

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Basic information

Entry
Database: EMDB / ID: EMD-22844
TitleCryo-electron microscopy structure of the heavy metal efflux pump CusA in a homogeneous binding copper(1) state
Map data
SampleCusA in symmetric copper binding form:
Cation efflux system protein CusA / ligand
Function / homology
Function and homology information


silver ion transmembrane transporter activity / silver ion transport / plasma membrane copper ion transport / response to silver ion / copper ion export / copper ion transmembrane transporter activity / detoxification of copper ion / xenobiotic transmembrane transporter activity / response to copper ion / cellular copper ion homeostasis ...silver ion transmembrane transporter activity / silver ion transport / plasma membrane copper ion transport / response to silver ion / copper ion export / copper ion transmembrane transporter activity / detoxification of copper ion / xenobiotic transmembrane transporter activity / response to copper ion / cellular copper ion homeostasis / copper ion binding / integral component of membrane / plasma membrane
Multidrug efflux transporter AcrB TolC docking domain, DN/DC subdomains / Cation efflux system CzcA/CusA/SilA/NccA/HelA/CnrA / Acriflavin resistance protein
Cation efflux system protein CusA
Biological speciesEscherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.4 Å
AuthorsMoseng MA
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)R01 AI145069 United States
CitationJournal: mBio / Year: 2021
Title: Cryo-EM Structures of CusA Reveal a Mechanism of Metal-Ion Export.
Authors: Mitchell A Moseng / Meinan Lyu / Tanadet Pipatpolkai / Przemyslaw Glaza / Corey C Emerson / Phoebe L Stewart / Phillip J Stansfeld / Edward W Yu /
Abstract: Gram-negative bacteria utilize the resistance-nodulation-cell division (RND) superfamily of efflux pumps to expel a variety of toxic compounds from the cell. The CusA membrane protein, which ...Gram-negative bacteria utilize the resistance-nodulation-cell division (RND) superfamily of efflux pumps to expel a variety of toxic compounds from the cell. The CusA membrane protein, which recognizes and extrudes biocidal Cu(I) and Ag(I) ions, belongs to the heavy-metal efflux (HME) subfamily of RND efflux pumps. We here report four structures of the trimeric CusA heavy-metal efflux pump in the presence of Cu(I) using single-particle cryo-electron microscopy (cryo-EM). We discover that different CusA protomers within the trimer are able to bind Cu(I) ions simultaneously. Our structural data combined with molecular dynamics (MD) simulations allow us to propose a mechanism for ion transport where each CusA protomer functions independently within the trimer. The bacterial RND superfamily of efflux pumps mediate resistance to a variety of biocides, including Cu(I) and Ag(I) ions. Here we report four cryo-EM structures of the trimeric CusA pump in the presence of Cu(I). Combined with MD simulations, our data indicate that each CusA protomer within the trimer recognizes and extrudes Cu(I) independently.
Validation ReportSummary, Full report, XML, About validation report
History
DepositionOct 13, 2020-
Header (metadata) releaseApr 14, 2021-
Map releaseApr 14, 2021-
UpdateApr 14, 2021-
Current statusApr 14, 2021Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.2
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.2
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7kf6
  • Surface level: 0.2
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_22844.map.gz / Format: CCP4 / Size: 13.4 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesX (Sec.)Y (Row.)Z (Col.)
1.08 Å/pix.
x 153 pix.
= 165.24 Å
1.08 Å/pix.
x 148 pix.
= 159.84 Å
1.08 Å/pix.
x 155 pix.
= 167.4 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

generated in cubic-lattice coordinate

Voxel sizeX=Y=Z: 1.08 Å
Density
Contour LevelBy AUTHOR: 0.2 / Movie #1: 0.2
Minimum - Maximum-0.6980122 - 1.257363
Average (Standard dev.)1.7434556e-12 (±0.09549716)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderZYX
Origin112114105
Dimensions148155153
Spacing153148155
CellA: 165.24 Å / B: 159.84001 Å / C: 167.40001 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.081.081.08
M x/y/z153148155
origin x/y/z0.0000.0000.000
length x/y/z165.240159.840167.400
α/β/γ90.00090.00090.000
start NX/NY/NZ105112114
NX/NY/NZ153148155
MAP C/R/S321
start NC/NR/NS114112105
NC/NR/NS155148153
D min/max/mean-0.6981.2570.000

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Supplemental data

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Sample components

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Entire CusA in symmetric copper binding form

EntireName: CusA in symmetric copper binding form / Number of components: 3

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Component #1: protein, CusA in symmetric copper binding form

ProteinName: CusA in symmetric copper binding form / Recombinant expression: No
SourceSpecies: Escherichia coli (E. coli)
Source (engineered)Expression System: Escherichia coli (E. coli)

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Component #2: protein, Cation efflux system protein CusA

ProteinName: Cation efflux system protein CusA / Number of Copies: 3 / Recombinant expression: No
MassTheoretical: 115.833945 kDa
SourceSpecies: Escherichia coli (E. coli)
Source (engineered)Expression System: Escherichia coli (E. coli)

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Component #3: ligand, COPPER (I) ION

LigandName: COPPER (I) ION / Number of Copies: 3 / Recombinant expression: No
MassTheoretical: 6.35459999999999905 MDa

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Experimental details

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Sample preparation

SpecimenSpecimen state: Particle / Method: cryo EM
Sample solutionpH: 7
VitrificationCryogen name: ETHANE / Temperature: 277.5 K / Humidity: 100 %

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 40 e/Å2 / Illumination mode: OTHER
LensImaging mode: BRIGHT FIELD
Specimen HolderModel: OTHER
CameraDetector: GATAN K2 SUMMIT (4k x 4k)

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Image processing

ProcessingMethod: single particle reconstruction / Number of projections: 3452
3D reconstructionResolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF

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Atomic model buiding

Modeling #1Refinement protocol: rigid body / Refinement space: REAL
Output model

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