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- EMDB-21356: Structure of a bacterial Atm1-family ABC exporter with MgADPVO4 bound -

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Basic information

Entry
Database: EMDB / ID: EMD-21356
TitleStructure of a bacterial Atm1-family ABC exporter with MgADPVO4 bound
Map data
Samplehomodimeric ATM1-type heavy metal exporter:
ATM1-type heavy metal exporter / (ligand) x 3
Function / homology
Function and homology information


response to mercury ion / ATPase-coupled transmembrane transporter activity / ion transport / ATPase activity / integral component of membrane / ATP binding / plasma membrane
AAA+ ATPase domain / ABC transporter-like / ABC transporter type 1, transmembrane domain / ABC transporter, conserved site / P-loop containing nucleoside triphosphate hydrolase / ABC transporter type 1, transmembrane domain superfamily / Type I protein exporter
ATM1-type heavy metal exporter
Biological speciesNovosphingobium aromaticivorans DSM 12444 (bacteria) / Novosphingobium aromaticivorans (strain ATCC 700278 / DSM 12444 / CIP 105152 / NBRC 16084 / F199) (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.03 Å
AuthorsFan C / Rees DC
Funding support United States, 1 items
OrganizationGrant numberCountry
Howard Hughes Medical Institute (HHMI) United States
CitationJournal: Proc. Natl. Acad. Sci. U.S.A. / Year: 2020
Title: A structural framework for unidirectional transport by a bacterial ABC exporter.
Authors: Chengcheng Fan / Jens T Kaiser / Douglas C Rees /
Abstract: The ATP-binding cassette (ABC) transporter of mitochondria (Atm1) mediates iron homeostasis in eukaryotes, while the prokaryotic homolog from (Atm1) can export glutathione derivatives and confer ...The ATP-binding cassette (ABC) transporter of mitochondria (Atm1) mediates iron homeostasis in eukaryotes, while the prokaryotic homolog from (Atm1) can export glutathione derivatives and confer protection against heavy-metal toxicity. To establish the structural framework underlying the Atm1 transport mechanism, we determined eight structures by X-ray crystallography and single-particle cryo-electron microscopy in distinct conformational states, stabilized by individual disulfide crosslinks and nucleotides. As Atm1 progresses through the transport cycle, conformational changes in transmembrane helix 6 (TM6) alter the glutathione-binding site and the associated substrate-binding cavity. Significantly, kinking of TM6 in the post-ATP hydrolysis state stabilized by MgADPVO eliminates this cavity, precluding uptake of glutathione derivatives. The presence of this cavity during the transition from the inward-facing to outward-facing conformational states, and its absence in the reverse direction, thereby provide an elegant and conceptually simple mechanism for enforcing the export directionality of transport by Atm1. One of the disulfide crosslinked Atm1 variants characterized in this work retains significant glutathione transport activity, suggesting that ATP hydrolysis and substrate transport by Atm1 may involve a limited set of conformational states with minimal separation of the nucleotide-binding domains in the inward-facing conformation.
Validation ReportPDB-ID: 6vqt

SummaryFull reportAbout validation report
History
DepositionFeb 5, 2020-
Header (metadata) releaseJul 29, 2020-
Map releaseJul 29, 2020-
UpdateJul 29, 2020-
Current statusJul 29, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.1
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.1
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6vqt
  • Surface level: 0.1
  • 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_21356.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.85 Å/pix.
x 300 pix.
= 255.66 Å
0.85 Å/pix.
x 300 pix.
= 255.66 Å
0.85 Å/pix.
x 300 pix.
= 255.66 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.8522 Å
Density
Contour LevelBy AUTHOR: 0.1 / Movie #1: 0.1
Minimum - Maximum-0.5005272 - 0.4887093
Average (Standard dev.)0.0009802915 (±0.06018416)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 255.65999 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.85220.85220.8522
M x/y/z300300300
origin x/y/z0.0000.0000.000
length x/y/z255.660255.660255.660
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ400400400
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS300300300
D min/max/mean-0.0430.3360.001

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Supplemental data

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Additional map: Sharpened map

Fileemd_21356_additional.map
AnnotationSharpened map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Half map: #2

Fileemd_21356_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Half map: #1

Fileemd_21356_half_map_2.map
Projections & Slices
AxesZYX

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Sample components

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Entire homodimeric ATM1-type heavy metal exporter

EntireName: homodimeric ATM1-type heavy metal exporter / Number of components: 5

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Component #1: protein, homodimeric ATM1-type heavy metal exporter

ProteinName: homodimeric ATM1-type heavy metal exporter / Recombinant expression: No
MassTheoretical: 133 kDa
SourceSpecies: Novosphingobium aromaticivorans DSM 12444 (bacteria)
Source (engineered)Expression System: Escherichia coli BL21(DE3) (bacteria)

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Component #2: protein, ATM1-type heavy metal exporter

ProteinName: ATM1-type heavy metal exporter / Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 67.771602 kDa
SourceSpecies: Novosphingobium aromaticivorans (strain ATCC 700278 / DSM 12444 / CIP 105152 / NBRC 16084 / F199) (bacteria)
Strain: ATCC 700278 / DSM 12444 / CIP 105152 / NBRC 16084 / F199
Source (engineered)Expression System: Escherichia coli BL21(DE3) (bacteria)

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Component #3: ligand, ADENOSINE-5'-DIPHOSPHATE

LigandName: ADENOSINE-5'-DIPHOSPHATE / Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 0.427201 kDa

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Component #4: ligand, VANADATE ION

LigandName: VANADATE ION / Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 0.114939 kDa

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Component #5: ligand, MAGNESIUM ION

LigandName: MAGNESIUM ION / Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 2.430505 MDa

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Experimental details

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Sample preparation

SpecimenSpecimen state: Particle / Method: cryo EM
Sample solutionSpecimen conc.: 1 mg/mL / pH: 7.5
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Temperature: 298 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: 48.6 e/Å2 / Illumination mode: FLOOD BEAM
LensMagnification: 29000 X (nominal) / Cs: 2.7 mm / Imaging mode: BRIGHT FIELD / Defocus: 1700.0 - 2400.0 nm
Specimen HolderModel: FEI TITAN KRIOS AUTOGRID HOLDER
CameraDetector: OTHER

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C2 (2 fold cyclic) / Number of projections: 169278
3D reconstructionSoftware: cryoSPARC / Resolution: 3.03 Å / Resolution method: FSC 0.143 CUT-OFF

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Atomic model buiding

Modeling #1Refinement protocol: rigid body / Refinement space: REAL
Input PDB model: 6PAR
Chain ID: A

Overall bvalue: 86
Output model

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