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- EMDB-20924: Single particle cryo-EM structure of KvAP -

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Basic information

Entry
Database: EMDB / ID: EMD-20924
TitleSingle particle cryo-EM structure of KvAP
Map data
SampleKvAP-6E1 Fab complex:
KvAP / 6E1 Fab / Voltage-gated potassium channel
Function / homologyIon transport domain / Voltage-gated potassium channel / voltage-gated potassium channel activity / regulation of ion transmembrane transport / voltage-gated potassium channel complex / identical protein binding / Voltage-gated potassium channel
Function and homology information
Biological speciesAeropyrum pernix (archaea) / unidentified (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 5.9 Å
AuthorsTao X / MacKinnon R
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical SciencesGM43949 United States
CitationJournal: Elife / Year: 2019
Title: Cryo-EM structure of the KvAP channel reveals a non-domain-swapped voltage sensor topology.
Authors: Xiao Tao / Roderick MacKinnon /
Abstract: Conductance in voltage-gated ion channels is regulated by membrane voltage through structural domains known as voltage sensors. A single structural class of voltage sensor domain exists, but two ...Conductance in voltage-gated ion channels is regulated by membrane voltage through structural domains known as voltage sensors. A single structural class of voltage sensor domain exists, but two different modes of voltage sensor attachment to the pore occur in nature: domain-swapped and non-domain-swapped. Since the more thoroughly studied Kv1-7, Nav and Cav channels have domain-swapped voltage sensors, much less is known about non-domain-swapped voltage-gated ion channels. In this paper, using cryo-EM, we show that KvAP from has non-domain-swapped voltage sensors as well as other unusual features. The new structure, together with previous functional data, suggests that KvAP and the Shaker channel, to which KvAP is most often compared, probably undergo rather different voltage-dependent conformational changes when they open.
Validation ReportPDB-ID: 6uwm

SummaryFull reportAbout validation report
History
DepositionNov 5, 2019-
Header (metadata) releaseDec 4, 2019-
Map releaseDec 4, 2019-
UpdateDec 4, 2019-
Current statusDec 4, 2019Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 6.07
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 6.07
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: : PDB-6uwm
  • Surface level: 6.07
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic models: PDB-6uwm
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_20924.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.)
1.03 Å/pix.
x 300 pix.
= 308.4 Å
1.03 Å/pix.
x 300 pix.
= 308.4 Å
1.03 Å/pix.
x 300 pix.
= 308.4 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.028 Å
Density
Contour LevelBy AUTHOR: 6.07 / Movie #1: 6.07
Minimum - Maximum-16.51102 - 26.106129
Average (Standard dev.)-0.22258796 (±0.9665328)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 308.4 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.0281.0281.028
M x/y/z300300300
origin x/y/z0.0000.0000.000
length x/y/z308.400308.400308.400
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ300300300
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS300300300
D min/max/mean-16.51126.106-0.223

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Supplemental data

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Sample components

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Entire KvAP-6E1 Fab complex

EntireName: KvAP-6E1 Fab complex / Number of components: 4

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Component #1: protein, KvAP-6E1 Fab complex

ProteinName: KvAP-6E1 Fab complex / Recombinant expression: No
MassTheoretical: 200 kDa

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Component #2: protein, KvAP

ProteinName: KvAP / Details: KvAP tetramer / Recombinant expression: No
MassTheoretical: 130 kDa
SourceSpecies: Aeropyrum pernix (archaea)
Source (engineered)Expression System: Escherichia coli BL21(DE3) (bacteria) / Vector: pET28a

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Component #3: protein, 6E1 Fab

ProteinName: 6E1 Fab / Recombinant expression: No
SourceSpecies: unidentified (others)

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Component #4: protein, Voltage-gated potassium channel

ProteinName: Voltage-gated potassium channel / Number of Copies: 4 / Recombinant expression: No
MassTheoretical: 31.649207 kDa
SourceSpecies: Aeropyrum pernix (archaea)
Source (engineered)Expression System: Escherichia coli BL21(DE3) (bacteria)

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Experimental details

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Sample preparation

SpecimenSpecimen state: Particle / Method: cryo EM
Sample solutionSpecimen conc.: 6 mg/mL / pH: 8
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Temperature: 295 K / Humidity: 100 % / Details: Blot for 4 seconds before plunging..

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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: 75 e/Å2 / Illumination mode: FLOOD BEAM
LensMagnification: 29000.0 X (nominal) / Cs: 2.7 mm / Imaging mode: BRIGHT FIELD / Defocus: 800.0 - 2200.0 nm
Specimen HolderModel: FEI TITAN KRIOS AUTOGRID HOLDER
CameraDetector: GATAN K2 SUMMIT (4k x 4k)

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Image acquisition

Image acquisitionNumber of digital images: 8000

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C4 (4 fold cyclic) / Number of projections: 67000
3D reconstructionSoftware: FREALIGN / Resolution: 5.9 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: 1ORS, 1ORQ
Output model

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