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- EMDB-19434: Cryo-EM structure of the light-driven sodium-pumping rhodopsin KR2 -

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Basic information

Entry
Database: EMDB / ID: EMD-19434
TitleCryo-EM structure of the light-driven sodium-pumping rhodopsin KR2
Map data
Sample
  • Complex: Sodium-pumping rhodopsin
    • Protein or peptide: Sodium pumping rhodopsin
  • Ligand: (2R)-2,3-dihydroxypropyl (9Z)-octadec-9-enoate
  • Ligand: EICOSANE
  • Ligand: RETINAL
  • Ligand: SODIUM ION
  • Ligand: OLEIC ACID
  • Ligand: water
Keywordsretinal / ion transport / sodium / rhodopsin / photocycle / pentamer / MEMBRANE PROTEIN
Function / homologyBacteriorhodopsin-like protein / Archaeal/bacterial/fungal rhodopsins / Bacteriorhodopsin-like protein / membrane / Sodium pumping rhodopsin
Function and homology information
Biological speciesDokdonia eikasta (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.32 Å
AuthorsKovalev K / Linares R
Funding support Germany, 1 items
OrganizationGrant numberCountry
EIPOD fellowship under Marie Sklodowska-Curie Actions COFUND847543 Germany
CitationJournal: To Be Published
Title: EasyGrid: A versatile platform for automated cryo-EM sample preparation and quality control
Authors: Gemin O / Armijo V / Papp G
History
DepositionJan 17, 2024-
Header (metadata) releaseJan 29, 2025-
Map releaseJan 29, 2025-
UpdateJan 29, 2025-
Current statusJan 29, 2025Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_19434.png

Downloads & links

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Map

FileDownload / File: emd_19434.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.84 Å/pix.
x 320 pix.
= 268.48 Å		0.84 Å/pix.
x 320 pix.
= 268.48 Å		0.84 Å/pix.
x 320 pix.
= 268.48 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.839 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.4
Minimum - Maximum-1.2463374 - 2.2708523
Average (Standard dev.)0.0014914346 (±0.06547618)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 268.47998 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: unsharpened map

Fileemd_19434_additional_1.map
Annotationunsharpened map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: mask for the refinement

Fileemd_19434_additional_2.map
Annotationmask for the refinement
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_19434_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_19434_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Sodium-pumping rhodopsin

EntireName: Sodium-pumping rhodopsin
Components
  • Complex: Sodium-pumping rhodopsin
    • Protein or peptide: Sodium pumping rhodopsin
  • Ligand: (2R)-2,3-dihydroxypropyl (9Z)-octadec-9-enoate
  • Ligand: EICOSANE
  • Ligand: RETINAL
  • Ligand: SODIUM ION
  • Ligand: OLEIC ACID
  • Ligand: water

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Supramolecule #1: Sodium-pumping rhodopsin

SupramoleculeName: Sodium-pumping rhodopsin / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1 / Details: KR2 rhodopsin solubilized in DDM
Source (natural)Organism: Dokdonia eikasta (bacteria)
Molecular weightTheoretical: 150 KDa

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Macromolecule #1: Sodium pumping rhodopsin

MacromoleculeName: Sodium pumping rhodopsin / type: protein_or_peptide / ID: 1 / Number of copies: 5 / Enantiomer: LEVO
Source (natural)Organism: Dokdonia eikasta (bacteria)
Molecular weightTheoretical: 31.540439 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MTQELGNANF ENFIGATEGF SEIAYQFTSH ILTLGYAVML AGLLYFILTI KNVDKKFQMS NILSAVVMVS AFLLLYAQAQ NWTSSFTFN EEVGRYFLDP SGDLFNNGYR YLNWLIDVPM LLFQILFVVS LTTSKFSSVR NQFWFSGAMM IITGYIGQFY E VSNLTAFL ...String:
MTQELGNANF ENFIGATEGF SEIAYQFTSH ILTLGYAVML AGLLYFILTI KNVDKKFQMS NILSAVVMVS AFLLLYAQAQ NWTSSFTFN EEVGRYFLDP SGDLFNNGYR YLNWLIDVPM LLFQILFVVS LTTSKFSSVR NQFWFSGAMM IITGYIGQFY E VSNLTAFL VWGAISSAFF FHILWVMKKV INEGKEGISP AGQKILSNIW ILFLISWTLY PGAYLMPYLT GVDGFLYSED GV MARQLVY TIADVSSKVI YGVLLGNLAI TLSKNKELVE ANS

UniProtKB: Sodium pumping rhodopsin

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Macromolecule #2: (2R)-2,3-dihydroxypropyl (9Z)-octadec-9-enoate

MacromoleculeName: (2R)-2,3-dihydroxypropyl (9Z)-octadec-9-enoate / type: ligand / ID: 2 / Number of copies: 5 / Formula: OLC
Molecular weightTheoretical: 356.54 Da
Chemical component information

ChemComp-OLC:
(2R)-2,3-dihydroxypropyl (9Z)-octadec-9-enoate

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Macromolecule #3: EICOSANE

MacromoleculeName: EICOSANE / type: ligand / ID: 3 / Number of copies: 66 / Formula: LFA
Molecular weightTheoretical: 282.547 Da
Chemical component information

ChemComp-LFA:
EICOSANE

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Macromolecule #4: RETINAL

MacromoleculeName: RETINAL / type: ligand / ID: 4 / Number of copies: 5 / Formula: RET
Molecular weightTheoretical: 284.436 Da
Chemical component information

ChemComp-RET:
RETINAL

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Macromolecule #5: SODIUM ION

MacromoleculeName: SODIUM ION / type: ligand / ID: 5 / Number of copies: 5
Molecular weightTheoretical: 22.99 Da

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Macromolecule #6: OLEIC ACID

MacromoleculeName: OLEIC ACID / type: ligand / ID: 6 / Number of copies: 4 / Formula: OLA
Molecular weightTheoretical: 282.461 Da
Chemical component information

ChemComp-OLA:
OLEIC ACID

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Macromolecule #7: water

MacromoleculeName: water / type: ligand / ID: 7 / Number of copies: 352 / 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

Concentration10 mg/mL
BufferpH: 8
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY ARRAY / Pretreatment - Type: PLASMA CLEANING / Pretreatment - Time: 10 sec. / Pretreatment - Atmosphere: AIR
Details: copper grids were first plasma-treated 15 times in EasyGrid,
VitrificationCryogen name: ETHANE / Chamber humidity: 85 % / Chamber temperature: 297 K / Details: Jet vitrified with EasyGrid.

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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: 6671 / Average exposure time: 1.6 sec. / Average electron dose: 40.3 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 70.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.4 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 105000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 1916000
Details: A first round of blob picking was performed and used to obtain an initial model. this initial model was used to generate templates that were then used to performed template picking, ...Details: A first round of blob picking was performed and used to obtain an initial model. this initial model was used to generate templates that were then used to performed template picking, resulting in 1916000 particles
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

6yc3

Final reconstructionApplied symmetry - Point group: C5 (5 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 2.32 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.2) / Number images used: 333078
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.2)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.2)
Final 3D classificationNumber classes: 100 / Avg.num./class: 3000 / Software - Name: cryoSPARC (ver. 4.2)
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-8rq5:
Cryo-EM structure of the light-driven sodium-pumping rhodopsin KR2

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