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- EMDB-49231: Open conformation of ArsA from L. ferriphilum in complex with MgA... -

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Basic information

Entry
Database: EMDB / ID: EMD-49231
TitleOpen conformation of ArsA from L. ferriphilum in complex with MgADP determined in the presence of arsenite
Map data
Sample
  • Complex: ArsA open state in complex with MgADP
    • Protein or peptide: Arsenite transporter ATPase-like protein,arsA
  • Ligand: ADENOSINE-5'-DIPHOSPHATE
  • Ligand: MAGNESIUM ION
KeywordsATPase / open conformation / arsenite / arsenic / ADP / Intradimeric Walker A / HYDROLASE
Function / homology
Function and homology information


arsenite-transporting ATPase / ATPase-coupled arsenite transmembrane transporter activity / ATP hydrolysis activity / ATP binding
Similarity search - Function
Arsenical pump-driving ATPase / Arsenical pump ATPase, ArsA/GET3 / Anion-transporting ATPase-like domain / Anion-transporting ATPase / ATPases associated with a variety of cellular activities / AAA+ ATPase domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
arsenite-transporting ATPase
Similarity search - Component
Biological speciesLeptospirillum ferriphilum (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.4 Å
AuthorsMahajan S / Rees DC / Clemons WM
Funding support United States, 1 items
OrganizationGrant numberCountry
Howard Hughes Medical Institute (HHMI) United States
CitationJournal: Proc Natl Acad Sci U S A / Year: 2025
Title: Nucleotide- and metalloid-driven conformational changes in the arsenite efflux ATPase ArsA.
Authors: Shivansh Mahajan / Ashley E Pall / Yancheng E Li / Timothy L Stemmler / Douglas C Rees / William M Clemons /
Abstract: Arsenite (As) is toxic to all organisms due to its ability to tightly bind exposed thiols within cells. An important As resistance mechanism in prokaryotes involves proteins encoded by the operon. A ...Arsenite (As) is toxic to all organisms due to its ability to tightly bind exposed thiols within cells. An important As resistance mechanism in prokaryotes involves proteins encoded by the operon. A central component of the operon in many bacteria is the cytoplasmic ATPase, ArsA, which orchestrates a series of nucleotide-dependent handoffs, starting with the capture of As by the ArsD metallochaperone and culminating in its removal from the cell by the ArsB efflux pump. Although the mechanism of ArsA has been widely studied, the molecular details of how nucleotide hydrolysis modulates these events remain unclear. ArsA is an archetypal member of the intradimeric Walker A (IWA) family of ATPases, implicated in a diversity of complex biological functions. Conformational changes typical of IWA ATPases have been postulated to drive these molecular events but have not been demonstrated. We report cryogenic electron microscopy (cryo-EM) structures of ArsA in MgADP-bound and MgATP-bound states, as well as a distinct MgATP-bound state liganded to As. X-ray absorption spectroscopy (XAS) confirmed three-coordinate binding of As to the conserved cysteines at the metalloid-binding site of the closed state. Coupled with biochemical characterization, our cryo-EM structures reveal key conformational changes in the ArsA catalytic cycle consistent with other IWA ATPases and provide the structural basis for allosteric activation of nucleotide hydrolysis by As. This work establishes how the nucleotide state of ArsA transiently creates a high-affinity binding site that can sequester metalloid within the cell, followed by a nucleotide-driven handoff to ArsB for efflux.
History
DepositionFeb 14, 2025-
Header (metadata) releaseAug 20, 2025-
Map releaseAug 20, 2025-
UpdateSep 10, 2025-
Current statusSep 10, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_49231.png

Downloads & links

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Map

FileDownload / File: emd_49231.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
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Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.65 Å/pix.
x 320 pix.
= 208. Å		0.65 Å/pix.
x 320 pix.
= 208. Å		0.65 Å/pix.
x 320 pix.
= 208. Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.65 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.09
Minimum - Maximum-0.37187928 - 0.53461504
Average (Standard dev.)0.000010890163 (±0.010558119)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 208.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: raw unsharpened map

Fileemd_49231_additional_1.map
Annotationraw unsharpened map
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Half map: #2

Fileemd_49231_half_map_1.map
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Half map: #1

Fileemd_49231_half_map_2.map
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Sample components

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Entire : ArsA open state in complex with MgADP

EntireName: ArsA open state in complex with MgADP
Components
  • Complex: ArsA open state in complex with MgADP
    • Protein or peptide: Arsenite transporter ATPase-like protein,arsA
  • Ligand: ADENOSINE-5'-DIPHOSPHATE
  • Ligand: MAGNESIUM ION

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Supramolecule #1: ArsA open state in complex with MgADP

SupramoleculeName: ArsA open state in complex with MgADP / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Details: ArsA incubated with 2 mM each of MgCl2 and ADP and 5 mM of sodium arsenite for 3 hours
Source (natural)Organism: Leptospirillum ferriphilum (bacteria) / Strain: ML-04

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Macromolecule #1: Arsenite transporter ATPase-like protein,arsA

MacromoleculeName: Arsenite transporter ATPase-like protein,arsA / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Leptospirillum ferriphilum (bacteria) / Strain: ML-04
Molecular weightTheoretical: 63.972004 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MGMKFLQLPP RFMFFTGKGG VGKTSIACAT SIQLANAGKR VLLVSTDPAS NVGQVFGVDI GNRVTPIPAV PHLSALEIDP EAAASAYRE RLVGPVRGVL PDDVVKGIEE SLSGACTTEI AAFDEFTALL TNAVLTADYQ HIIFDTAPTG HTIRLLQLPG A WSGFLEAG ...String:
MGMKFLQLPP RFMFFTGKGG VGKTSIACAT SIQLANAGKR VLLVSTDPAS NVGQVFGVDI GNRVTPIPAV PHLSALEIDP EAAASAYRE RLVGPVRGVL PDDVVKGIEE SLSGACTTEI AAFDEFTALL TNAVLTADYQ HIIFDTAPTG HTIRLLQLPG A WSGFLEAG KGDASCLGPL AGLEKQRTQY KAAVEALADP LQTRLVLVAR AQQATLREVA RTHEELATIG IKQQHLVING IL PSAEAAN DPLAAAIHER EQTALKNIPA TLTSLPRDLV QLKPFNLVGL DALRQLLTDL PLQAHVAADA PIELDEPGMG DLV DGIEAD GHGLVMLMGK GGVGKTTLAA AIAVELAHRG LPVHLTTSDP AAHLTDTLEA SLDNLTVSRI DPHAETERYR QHVL ETKGA QLDAEGRALL EEDLHSPCTE EIAVFQAFSR IIREAGKKFV VMDTAPTGHT LLLLDATGAY HREVRRQMGN KGTHF TTPM MQLRDPNQTK VLVVTLAETT PVLEAAKLQA DLRRAGIEPW AWIINTSVAA ASAKSPLLRQ RAANELREIN AVANHH ADR YAVVPLLKEE PIGAERLRAL IHPQTHHHHH H

UniProtKB: arsenite-transporting ATPase

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Macromolecule #2: ADENOSINE-5'-DIPHOSPHATE

MacromoleculeName: ADENOSINE-5'-DIPHOSPHATE / type: ligand / ID: 2 / Number of copies: 2 / Formula: ADP
Molecular weightTheoretical: 427.201 Da
Chemical component information

ChemComp-ADP:
ADENOSINE-5'-DIPHOSPHATE / ADP, energy-carrying molecule*YM

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Macromolecule #3: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 3 / Number of copies: 2 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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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

Concentration10 mg/mL
BufferpH: 7.5
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV
DetailsArsA incubated with 2 mM each of MgCl2 and ADP and 5 mM of sodium arsenite for 3 hours

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 70.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 130000
Sample stageCooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: NONE
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 103258
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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