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- EMDB-8414: Ca2+ bound aplysia Slo1 -

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Basic information

Entry
Database: EMDB / ID: EMD-8414
TitleCa2+ bound aplysia Slo1
Map dataAplysia Slo1 in 1 mM EDTA
Sample
  • Organelle or cellular component: Ca2+ bound aplysia Slo1
    • Protein or peptide: High conductance calcium-activated potassium channel
  • Ligand: (1R)-2-{[(S)-{[(2S)-2,3-dihydroxypropyl]oxy}(hydroxy)phosphoryl]oxy}-1-[(hexadecanoyloxy)methyl]ethyl (9Z)-octadec-9-enoate
Keywordsion channel / K+ channel / Ca2+ bound / high conductance / MEMBRANE PROTEIN
Function / homology
Function and homology information


large conductance calcium-activated potassium channel activity / postsynaptic membrane
Similarity search - Function
: / Ca2+-activated K+ channel Slowpoke, TrkA_C like domain / Calcium-activated potassium channel BK, alpha subunit / : / Calcium-activated BK potassium channel alpha subunit / Calcium-activated potassium channel slowpoke-like RCK domain / Voltage-dependent channel domain superfamily / Ion transport domain / Ion transport protein / NAD(P)-binding domain superfamily
Similarity search - Domain/homology
Biological speciesAplysia californica (California sea hare)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.8 Å
AuthorsMacKinnon R / Tao X
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM43949 United States
Howard Hughes Medical Institute (HHMI) United States
CitationJournal: Nature / Year: 2017
Title: Structural basis for gating the high-conductance Ca-activated K channel.
Authors: Richard K Hite / Xiao Tao / Roderick MacKinnon /
Abstract: The precise control of an ion channel gate by environmental stimuli is crucial for the fulfilment of its biological role. The gate in Slo1 K channels is regulated by two separate stimuli, ...The precise control of an ion channel gate by environmental stimuli is crucial for the fulfilment of its biological role. The gate in Slo1 K channels is regulated by two separate stimuli, intracellular Ca concentration and membrane voltage. Slo1 is thus central to understanding the relationship between intracellular Ca and membrane excitability. Here we present the Slo1 structure from Aplysia californica in the absence of Ca and compare it with the Ca-bound channel. We show that Ca binding at two unique binding sites per subunit stabilizes an expanded conformation of the Ca sensor gating ring. These conformational changes are propagated from the gating ring to the pore through covalent linkers and through protein interfaces formed between the gating ring and the voltage sensors. The gating ring and the voltage sensors are directly connected through these interfaces, which allow membrane voltage to regulate gating of the pore by influencing the Ca sensors.
History
DepositionOct 4, 2016-
Header (metadata) releaseNov 16, 2016-
Map releaseDec 28, 2016-
UpdateMar 13, 2024-
Current statusMar 13, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.11
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.11
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-5tji
  • Surface level: 0.11
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_8414.png

Downloads & links

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Map

FileDownload / File: emd_8414.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationAplysia Slo1 in 1 mM EDTA
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.35 Å/pix.
x 256 pix.
= 345.6 Å		1.35 Å/pix.
x 256 pix.
= 345.6 Å		1.35 Å/pix.
x 256 pix.
= 345.6 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.35 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.11 / Movie #1: 0.11
Minimum - Maximum-0.036365576 - 0.204226
Average (Standard dev.)0.0031882646 (±0.01626475)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 345.6 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.351.351.35
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z345.600345.600345.600
α/β/γ90.00090.00090.000
start NX/NY/NZ-152-37
NX/NY/NZ998271
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS256256256
D min/max/mean-0.0360.2040.003

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Supplemental data

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Sample components

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Entire : Ca2+ bound aplysia Slo1

EntireName: Ca2+ bound aplysia Slo1
Components
  • Organelle or cellular component: Ca2+ bound aplysia Slo1
    • Protein or peptide: High conductance calcium-activated potassium channel
  • Ligand: (1R)-2-{[(S)-{[(2S)-2,3-dihydroxypropyl]oxy}(hydroxy)phosphoryl]oxy}-1-[(hexadecanoyloxy)methyl]ethyl (9Z)-octadec-9-enoate

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Supramolecule #1: Ca2+ bound aplysia Slo1

SupramoleculeName: Ca2+ bound aplysia Slo1 / type: organelle_or_cellular_component / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Aplysia californica (California sea hare)
Molecular weightTheoretical: 400 KDa

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Macromolecule #1: High conductance calcium-activated potassium channel

MacromoleculeName: High conductance calcium-activated potassium channel / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Aplysia californica (California sea hare)
Molecular weightTheoretical: 120.308 KDa
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)
SequenceString: MASSSSTSCE PGDRQWYSFL ASSLVTFGSG LVVIIIYRIV LWLCCRKKKC IQVSNPVPTA RTTSLDQKSF MKNSDPEIGW MTEAKDWAG ELISGQTTTG RILVGLVFLL SIASLIIYFI DASTNTSVET CLPWSSSTTQ QVDLAFNVFF MIYFFIRFVA A NDKLWFWV ...String:
MASSSSTSCE PGDRQWYSFL ASSLVTFGSG LVVIIIYRIV LWLCCRKKKC IQVSNPVPTA RTTSLDQKSF MKNSDPEIGW MTEAKDWAG ELISGQTTTG RILVGLVFLL SIASLIIYFI DASTNTSVET CLPWSSSTTQ QVDLAFNVFF MIYFFIRFVA A NDKLWFWV ELFSFVDYFT IPPSFVAIYL DRNWLGLRFL RALRLMSIPD ILTYLNVLKT STLIRLVQLV VSFVSLWLTA AG FLHLLEN SGDPFFDFGN AQHLTYWECL YFLMVTMSTV GFGDIFATTV LGRTFVVIFI MIFIGLFASF IPEIAEILGK RQK YGGSYK KERGKRHVVV CGYITFDSVS NFLKDFLHKD REDVDVEIVF LHKGLPGLEL EGLLKRHFTQ VEYFWGSVMD ANDL ERVKI QEADACLVLA NKYCQDPDQE DAANIMRVIS IKNYHSDIKV IVQLLQYHNK AYLLNIPSWD WKRGDDAVCV AELKL GFIA QSCLAPGFST LMANLFTMRS YKPTPEMSQW QTDYMRGTGM EMYTEYLSSA FNALTFPEAA ELCFSKLKLL LLAIEV RQE DTRESTLAIN PGPKVKIENA TQGFFIAESA EEVKRAFYYC KNCHANVSDV RQIKKCKCRP LAMFKKGAAA VLALQRT PG LAVEPDGEAN DKDKSRGTST SKAVTSFPEK RKPQSRRKPS TTLKSKSPSE DSVPPPPPPV DEPRKFDSTG MFHWCPDR P LNDCLQDRSQ ASASGLRNHV VVCLFADAAS PLIGLRNLVM PLRASNFHYH ELKPTIIVGN LDYLHREWKT LQNFPKLSI LPGSPLNRAN LRAVNINLCD MCVIVSAKDR NMEDPNLVDK EAILCSLNIK AMTFDDTMGL IQSSNFVPGG FSPLHENKRS QAGANVPLI TELANDSNVQ FLDQDDDDDP DTELYMTQPF ACGTAFAVSV LDSLMSTSYF NDNALTLIRT LITGGATPEL E QILAEGAG MRGGYCSPAV LANRDRCRVA QISLFDGPLA QFGQGGHYGE LFVYALRHFG ILCIGLYRFR DTNESVRSPS SK RYVITNP PEDFPLLPTD QVYVLTYKQI TNH

UniProtKB: BK channel

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Macromolecule #2: (1R)-2-{[(S)-{[(2S)-2,3-dihydroxypropyl]oxy}(hydroxy)phosphoryl]o...

MacromoleculeName: (1R)-2-{[(S)-{[(2S)-2,3-dihydroxypropyl]oxy}(hydroxy)phosphoryl]oxy}-1-[(hexadecanoyloxy)methyl]ethyl (9Z)-octadec-9-enoate
type: ligand / ID: 2 / Number of copies: 1 / Formula: PGW
Molecular weightTheoretical: 749.007 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

Concentration7 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
20.0 mMTrisTris
450.0 mMKClKCl
20.0 mMDTTDTT
2.0 mMTCEPTCEP
0.025 %DDMDDM
0.005 %CHSCHS
1.0 mMEDTAEDTA
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 10 sec. / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 85 % / Chamber temperature: 293 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsSpherical aberration corrector: FEI Cs corrector on Titan Krios
Energy filter - Name: GIF / Energy filter - Lower energy threshold: -15 eV / Energy filter - Upper energy threshold: 15 eV
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Digitization - Dimensions - Width: 7420 pixel / Digitization - Dimensions - Height: 7676 pixel / Digitization - Frames/image: 1-50 / Number grids imaged: 1 / Number real images: 4000 / Average exposure time: 0.3 sec. / Average electron dose: 1.4 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 22500
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: 638000
Startup modelType of model: OTHER
Details: An initial model generated from 2D class averages in EMAN2
Final reconstructionApplied symmetry - Point group: C4 (4 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 3.8 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: FREALIGN / Number images used: 60000
Initial angle assignmentType: PROJECTION MATCHING / Software - Name: RELION
Final angle assignmentType: PROJECTION MATCHING / Software - Name: FREALIGN
Final 3D classificationNumber classes: 6 / Avg.num./class: 75000 / Software - Name: RELION

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Atomic model buiding 1

RefinementSpace: RECIPROCAL / Protocol: AB INITIO MODEL / Overall B value: 160
Output model

PDB-5tji:
Ca2+ bound aplysia Slo1

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