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- EMDB-27135: hBest1 Ca2+-unbound closed state -

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Basic information

Entry
Database: EMDB / ID: EMD-27135
TitlehBest1 Ca2+-unbound closed state
Map data
Sample
  • Complex: hBest1 Ca2+-unbound closed state
    • Protein or peptide: Bestrophin-1
KeywordsIon channel / chloride channel / anion channel / pentamer / TRANSPORT PROTEIN
Function / homology
Function and homology information


membrane microdomain / bicarbonate channel activity / transepithelial chloride transport / gamma-aminobutyric acid secretion, neurotransmission / detection of light stimulus involved in visual perception / ligand-gated channel activity / intracellularly calcium-gated chloride channel activity / glutamate secretion / bicarbonate transmembrane transporter activity / chloride transport ...membrane microdomain / bicarbonate channel activity / transepithelial chloride transport / gamma-aminobutyric acid secretion, neurotransmission / detection of light stimulus involved in visual perception / ligand-gated channel activity / intracellularly calcium-gated chloride channel activity / glutamate secretion / bicarbonate transmembrane transporter activity / chloride transport / chloride channel activity / protein complex oligomerization / regulation of calcium ion transport / chloride channel complex / visual perception / basal plasma membrane / regulation of synaptic plasticity / Stimuli-sensing channels / presynapse / monoatomic ion transmembrane transport / basolateral plasma membrane / identical protein binding / membrane / plasma membrane / cytosol
Similarity search - Function
Bestrophin / Bestrophin/UPF0187 / Bestrophin, RFP-TM, chloride channel
Similarity search - Domain/homology
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.11 Å
AuthorsOwji AP / Kittredge A / Hendrickson WA / Tingting Y
Funding support United States, 3 items
OrganizationGrant numberCountry
National Institutes of Health/National Eye Institute (NIH/NEI)EY030763-03 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM107462-08 United States
National Institutes of Health/National Eye Institute (NIH/NEI)GM127652-06 United States
CitationJournal: Nat Commun / Year: 2022
Title: Structures and gating mechanisms of human bestrophin anion channels.
Authors: Aaron P Owji / Jiali Wang / Alec Kittredge / Zada Clark / Yu Zhang / Wayne A Hendrickson / Tingting Yang /
Abstract: Bestrophin-1 (Best1) and bestrophin-2 (Best2) are two members of the bestrophin family of calcium (Ca)-activated chloride (Cl) channels with critical involvement in ocular physiology and direct ...Bestrophin-1 (Best1) and bestrophin-2 (Best2) are two members of the bestrophin family of calcium (Ca)-activated chloride (Cl) channels with critical involvement in ocular physiology and direct pathological relevance. Here, we report cryo-EM structures of wild-type human Best1 and Best2 in various states at up to 1.8 Å resolution. Ca-bound Best1 structures illustrate partially open conformations at the two Ca-dependent gates of the channels, in contrast to the fully open conformations observed in Ca-bound Best2, which is in accord with the significantly smaller currents conducted by Best1 in electrophysiological recordings. Comparison of the closed and open states reveals a C-terminal auto-inhibitory segment (AS), which constricts the channel concentrically by wrapping around the channel periphery in an inter-protomer manner and must be released to allow channel opening. Our results demonstrate that removing the AS from Best1 and Best2 results in truncation mutants with similar activities, while swapping the AS between Best1 and Best2 results in chimeric mutants with swapped activities, underlying a key role of the AS in determining paralog specificity among bestrophins.
History
DepositionMay 27, 2022-
Header (metadata) releaseJul 13, 2022-
Map releaseJul 13, 2022-
UpdateFeb 14, 2024-
Current statusFeb 14, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_27135.png

Downloads & links

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Map

FileDownload / File: emd_27135.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.)
0.83 Å/pix.
x 300 pix.
= 249. Å		0.83 Å/pix.
x 300 pix.
= 249. Å		0.83 Å/pix.
x 300 pix.
= 249. Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.83 Å
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Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.5
Minimum - Maximum-1.5107213 - 2.6918924
Average (Standard dev.)0.004751266 (±0.09521075)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 249.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: full map

Fileemd_27135_additional_1.map
Annotationfull map
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Half map: half map 1

Fileemd_27135_half_map_1.map
Annotationhalf map 1
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Half map: half map 2

Fileemd_27135_half_map_2.map
Annotationhalf map 2
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Sample components

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Entire : hBest1 Ca2+-unbound closed state

EntireName: hBest1 Ca2+-unbound closed state
Components
  • Complex: hBest1 Ca2+-unbound closed state
    • Protein or peptide: Bestrophin-1

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Supramolecule #1: hBest1 Ca2+-unbound closed state

SupramoleculeName: hBest1 Ca2+-unbound closed state / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 338 KDa

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Macromolecule #1: Bestrophin-1

MacromoleculeName: Bestrophin-1 / type: protein_or_peptide / ID: 1 / Number of copies: 5 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 67.760469 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MTITYTSQVA NARLGSFSRL LLCWRGSIYK LLYGEFLIFL LCYYIIRFIY RLALTEEQQL MFEKLTLYCD SYIQLIPISF VLGFYVTLV VTRWWNQYEN LPWPDRLMSL VSGFVEGKDE QGRLLRRTLI RYANLGNVLI LRSVSTAVYK RFPSAQHLVQ A GFMTPAEH ...String:
MTITYTSQVA NARLGSFSRL LLCWRGSIYK LLYGEFLIFL LCYYIIRFIY RLALTEEQQL MFEKLTLYCD SYIQLIPISF VLGFYVTLV VTRWWNQYEN LPWPDRLMSL VSGFVEGKDE QGRLLRRTLI RYANLGNVLI LRSVSTAVYK RFPSAQHLVQ A GFMTPAEH KQLEKLSLPH NMFWVPWVWF ANLSMKAWLG GRIRDPILLQ SLLNEMNTLR TQCGHLYAYD WISIPLVYTQ VV TVAVYSF FLTCLVGRQF LNPAKAYPGH ELDLVVPVFT FLQFFFYVGW LKVAEQLINP FGEDDDDFET NWIVDRNLQV SLL AVDEMH QDLPRMEPDM YWNKPEPQPP YTAASAQFRR ASFMGSTFNI SLNKEEMEFQ PNQEDEEDAH AGIIGRFLGL QSHD HHPPR ANSRTKLLWP KRESLLHEGL PKNHKAAKQN VRGQEDNKAW KLKAVDAFKS APLYQRPGYY SAPQTPLSPT PMFFP LEPS APSKLHSVTG IDTKDKSLKT VSSGAKKSFE LLSESDGALM EHPEVSQVRR KTVEFNLTDM PEIPENHLKE PLEQSP TNI HTTLKDHMDP YWALENRDEA HS

UniProtKB: Bestrophin-1

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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

Concentration5 mg/mL
BufferpH: 7.8
Component:
ConcentrationFormulaName
200.0 mMNaClsodium chloride
40.0 mM(4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid)
0.005 %glyco-diosgenin
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 283.15 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeTFS KRIOS
Specialist opticsEnergy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number real images: 5558 / Average electron dose: 58.0 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: 2.0 µm / Nominal defocus min: 1.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 2079411
Startup modelType of model: OTHER / Details: Ab initio
Final reconstructionApplied symmetry - Point group: C5 (5 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 3.11 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 94625
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC

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