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- EMDB-18609: Cryo-EM structure of the light-driven sodium pump ErNaR in the pe... -

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Basic information

Entry
Database: EMDB / ID: EMD-18609
TitleCryo-EM structure of the light-driven sodium pump ErNaR in the pentameric form at pH 8.0
Map data
Sample
  • Complex: Light-driven sodium pump ErNaR
    • Protein or peptide: Bacteriorhodopsin-like protein
  • Ligand: EICOSANE
  • Ligand: DODECYL-BETA-D-MALTOSIDE
  • Ligand: water
Keywordsretinal / ion transport / rhodopsin / photocycle / sodium transport / MEMBRANE PROTEIN
Function / homologyBacteriorhodopsin-like protein / Archaeal/bacterial/fungal rhodopsins / Bacteriorhodopsin-like protein / membrane / Bacteriorhodopsin-like protein
Function and homology information
Biological speciesErythrobacter (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.63 Å
AuthorsKovalev K / Podoliak E / Lamm GHU / Marin E / Stetsenko A / Guskov A
Funding support Germany, 1 items
OrganizationGrant numberCountry
EIPOD fellowship under Marie Sklodowska-Curie Actions COFUND847543 Germany
CitationJournal: Nat Commun / Year: 2024
Title: A subgroup of light-driven sodium pumps with an additional Schiff base counterion.
Authors: E Podoliak / G H U Lamm / E Marin / A V Schellbach / D A Fedotov / A Stetsenko / M Asido / N Maliar / G Bourenkov / T Balandin / C Baeken / R Astashkin / T R Schneider / A Bateman / J ...Authors: E Podoliak / G H U Lamm / E Marin / A V Schellbach / D A Fedotov / A Stetsenko / M Asido / N Maliar / G Bourenkov / T Balandin / C Baeken / R Astashkin / T R Schneider / A Bateman / J Wachtveitl / I Schapiro / V Busskamp / A Guskov / V Gordeliy / A Alekseev / K Kovalev /
Abstract: Light-driven sodium pumps (NaRs) are unique ion-transporting microbial rhodopsins. The major group of NaRs is characterized by an NDQ motif and has two aspartic acid residues in the central region ...Light-driven sodium pumps (NaRs) are unique ion-transporting microbial rhodopsins. The major group of NaRs is characterized by an NDQ motif and has two aspartic acid residues in the central region essential for sodium transport. Here we identify a subgroup of the NDQ rhodopsins bearing an additional glutamic acid residue in the close vicinity to the retinal Schiff base. We thoroughly characterize a member of this subgroup, namely the protein ErNaR from Erythrobacter sp. HL-111 and show that the additional glutamic acid results in almost complete loss of pH sensitivity for sodium-pumping activity, which is in contrast to previously studied NaRs. ErNaR is capable of transporting sodium efficiently even at acidic pH levels. X-ray crystallography and single particle cryo-electron microscopy reveal that the additional glutamic acid residue mediates the connection between the other two Schiff base counterions and strongly interacts with the aspartic acid of the characteristic NDQ motif. Hence, it reduces its pKa. Our findings shed light on a subgroup of NaRs and might serve as a basis for their rational optimization for optogenetics.
History
DepositionOct 6, 2023-
Header (metadata) releaseApr 24, 2024-
Map releaseApr 24, 2024-
UpdateApr 24, 2024-
Current statusApr 24, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_18609.png

Downloads & links

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Map

FileDownload / File: emd_18609.map.gz / Format: CCP4 / Size: 1000 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
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AxesZ (Sec.)Y (Row.)X (Col.)
0.42 Å/pix.
x 640 pix.
= 267.52 Å		0.42 Å/pix.
x 640 pix.
= 267.52 Å		0.42 Å/pix.
x 640 pix.
= 267.52 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.418 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.2
Minimum - Maximum-0.64079964 - 0.96789604
Average (Standard dev.)0.00030385575 (±0.022942018)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions640640640
Spacing640640640
CellA=B=C: 267.52002 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_18609_half_map_1.map
Projections & Slices
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Half map: #2

Fileemd_18609_half_map_2.map
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Sample components

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Entire : Light-driven sodium pump ErNaR

EntireName: Light-driven sodium pump ErNaR
Components
  • Complex: Light-driven sodium pump ErNaR
    • Protein or peptide: Bacteriorhodopsin-like protein
  • Ligand: EICOSANE
  • Ligand: DODECYL-BETA-D-MALTOSIDE
  • Ligand: water

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Supramolecule #1: Light-driven sodium pump ErNaR

SupramoleculeName: Light-driven sodium pump ErNaR / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Erythrobacter (bacteria)
Molecular weightTheoretical: 36 KDa

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Macromolecule #1: Bacteriorhodopsin-like protein

MacromoleculeName: Bacteriorhodopsin-like protein / type: protein_or_peptide / ID: 1 / Number of copies: 5 / Enantiomer: LEVO
Source (natural)Organism: Erythrobacter (bacteria)
Molecular weightTheoretical: 32.046049 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: (FME)PSIENFLAY DFWQYDVIRH LFAFSTAVFL AGLVYFAMTA RTTAPNYRLS ANISAVVMVS AALELGQLWL LWNESF QWA ELQGSFVPVA GERFSNGYRY MNWLIDVPML ATQLVVVCGF VGTELRNRWA KLTIAGVLMI LTGYVGQYFE PAVAGVP GY EGAEQFWIWG ...String:
(FME)PSIENFLAY DFWQYDVIRH LFAFSTAVFL AGLVYFAMTA RTTAPNYRLS ANISAVVMVS AALELGQLWL LWNESF QWA ELQGSFVPVA GERFSNGYRY MNWLIDVPML ATQLVVVCGF VGTELRNRWA KLTIAGVLMI LTGYVGQYFE PAVAGVP GY EGAEQFWIWG IISTAFFVWM LLILANAVRN PQGAPSDEVR SRLKFCFWFL LATWSIYPFA YAMPLFAPTA DGVVVRQV I YTVADVSS(LYR)L VFGVILSQVA LRRSAEEGFE PARVASGEFD ERAPAR

UniProtKB: Bacteriorhodopsin-like protein

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Macromolecule #2: EICOSANE

MacromoleculeName: EICOSANE / type: ligand / ID: 2 / Number of copies: 46 / Formula: LFA
Molecular weightTheoretical: 282.547 Da
Chemical component information

ChemComp-LFA:
EICOSANE

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Macromolecule #3: DODECYL-BETA-D-MALTOSIDE

MacromoleculeName: DODECYL-BETA-D-MALTOSIDE / type: ligand / ID: 3 / Number of copies: 10 / Formula: LMT
Molecular weightTheoretical: 510.615 Da
Chemical component information

ChemComp-LMT:
DODECYL-BETA-D-MALTOSIDE / detergent*YM

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Macromolecule #4: water

MacromoleculeName: water / type: ligand / ID: 4 / Number of copies: 195 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
WATER

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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

Concentration7 mg/mL
BufferpH: 4.3 / Component: (Name: tris-HCl, sodium chloride)
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 300
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 293 K

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 1 / Number real images: 3865 / Average electron dose: 50.0 e/Å2 / Details: Counted super res mode
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: OTHER / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 105000
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Final reconstructionApplied symmetry - Point group: C5 (5 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 2.63 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.0.2) / Number images used: 437017
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.0.2)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.0.2)
Final 3D classificationNumber classes: 20 / Software - Name: cryoSPARC (ver. 4.0.2)

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