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- PDB-9kc9: Cryo-EM structure of docked mouse bestrophin-1 in a partial open state -

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Basic information

Entry
Database: PDB / ID: 9kc9
TitleCryo-EM structure of docked mouse bestrophin-1 in a partial open state
ComponentsBestrophin-1,Soluble cytochrome b562
KeywordsMEMBRANE PROTEIN / Docked / Partial open / Calcium-bound
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 / Stimuli-sensing channels / glutamate secretion / 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 / Stimuli-sensing channels / glutamate secretion / chloride transport / chloride channel activity / protein complex oligomerization / regulation of calcium ion transport / chloride channel complex / electron transport chain / regulation of synaptic plasticity / presynapse / basolateral plasma membrane / periplasmic space / electron transfer activity / iron ion binding / heme binding / metal ion binding / identical protein binding / plasma membrane
Similarity search - Function
Bestrophin / Bestrophin/UPF0187 / Bestrophin, RFP-TM, chloride channel / Cytochrome b562 / Cytochrome b562 / Cytochrome c/b562
Similarity search - Domain/homology
Bestrophin-1 / Soluble cytochrome b562
Similarity search - Component
Biological speciesMus musculus (house mouse)
Escherichia coli (E. coli)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.1 Å
AuthorsLim, H.H. / Kim, K.W. / Ko, A.
Funding support Korea, Republic Of, 2items
OrganizationGrant numberCountry
National Research Foundation (NRF, Korea) Korea, Republic Of
National Research Foundation (NRF, Korea)2020 M3E5D9079 to H.-H.L. Korea, Republic Of
Citation
Journal: Mol Cells / Year: 2025
Title: Cryo-EM structures of mouse bestrophin 1 channel in closed and partially open conformations.
Authors: Kwon-Woo Kim / Euna Lee / Ara Ko / Junmo Hwang / Kunwoong Park / Byoung-Cheol Lee / Ki Woo Kim / Won-Jong Oh / Kyuhyung Kim / Hyun-Ho Lim /
Abstract: Bestrophin 1 (BEST1) channels are calcium-activated Cl channels involved in diverse physiological processes, including gliotransmitter release in astrocytes. Although human and chicken BEST1 ...Bestrophin 1 (BEST1) channels are calcium-activated Cl channels involved in diverse physiological processes, including gliotransmitter release in astrocytes. Although human and chicken BEST1 orthologs have been extensively studied, the structural and functional properties of mouse BEST1 (mBEST1) remain poorly understood. In this study, we characterized the structure-function of mBEST1-BF, a C-terminally tagged variant, using whole-cell patch-clamp recordings, surface biotinylation assays, and single-particle cryo-electron microscopy. Cryo-electron microscopy structural analysis of mBEST1-BF revealed closed and partially open conformations. Comparative analysis with human and chicken BEST1 orthologs highlighted conserved calcium-binding and gating mechanisms, with distinct features in mBEST1, including a wider aperture sufficient to accommodate dehydrated Cl ions and potential anion-binding sites near Val205 and Gln208 residues. The disordered C-terminal region of mBEST1 remains unresolved, suggesting it may require stabilizing factors for structural determination. Additionally, the autoinhibitory domain, which includes Ser354, likely plays a key role in regulating gating, with Ser354 potentially serving as a phosphorylation site that modulates channel activity. Our findings provide structural and functional insights into mBEST1 and suggest mechanisms underlying its unique gating and ion permeation properties.
#1: Journal: Acta Crystallogr D Struct Biol / Year: 2019
Title: Macromolecular structure determination using X-rays, neutrons and electrons: recent developments in Phenix.
Authors: Dorothee Liebschner / Pavel V Afonine / Matthew L Baker / Gábor Bunkóczi / Vincent B Chen / Tristan I Croll / Bradley Hintze / Li Wei Hung / Swati Jain / Airlie J McCoy / Nigel W Moriarty ...Authors: Dorothee Liebschner / Pavel V Afonine / Matthew L Baker / Gábor Bunkóczi / Vincent B Chen / Tristan I Croll / Bradley Hintze / Li Wei Hung / Swati Jain / Airlie J McCoy / Nigel W Moriarty / Robert D Oeffner / Billy K Poon / Michael G Prisant / Randy J Read / Jane S Richardson / David C Richardson / Massimo D Sammito / Oleg V Sobolev / Duncan H Stockwell / Thomas C Terwilliger / Alexandre G Urzhumtsev / Lizbeth L Videau / Christopher J Williams / Paul D Adams /
Abstract: Diffraction (X-ray, neutron and electron) and electron cryo-microscopy are powerful methods to determine three-dimensional macromolecular structures, which are required to understand biological ...Diffraction (X-ray, neutron and electron) and electron cryo-microscopy are powerful methods to determine three-dimensional macromolecular structures, which are required to understand biological processes and to develop new therapeutics against diseases. The overall structure-solution workflow is similar for these techniques, but nuances exist because the properties of the reduced experimental data are different. Software tools for structure determination should therefore be tailored for each method. Phenix is a comprehensive software package for macromolecular structure determination that handles data from any of these techniques. Tasks performed with Phenix include data-quality assessment, map improvement, model building, the validation/rebuilding/refinement cycle and deposition. Each tool caters to the type of experimental data. The design of Phenix emphasizes the automation of procedures, where possible, to minimize repetitive and time-consuming manual tasks, while default parameters are chosen to encourage best practice. A graphical user interface provides access to many command-line features of Phenix and streamlines the transition between programs, project tracking and re-running of previous tasks.
History
DepositionNov 1, 2024Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Mar 19, 2025Provider: repository / Type: Initial release
Revision 1.1Apr 2, 2025Group: Data collection / Database references / Category: citation / em_admin / Item: _citation.journal_volume / _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Bestrophin-1,Soluble cytochrome b562
E: Bestrophin-1,Soluble cytochrome b562
B: Bestrophin-1,Soluble cytochrome b562
C: Bestrophin-1,Soluble cytochrome b562
D: Bestrophin-1,Soluble cytochrome b562
hetero molecules


Theoretical massNumber of molelcules
Total (without water)396,98120
Polymers396,4265
Non-polymers55515
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

#1: Protein
Bestrophin-1,Soluble cytochrome b562 / Vitelliform macular dystrophy protein 2 homolog / Cytochrome b-562


Mass: 79285.266 Da / Num. of mol.: 5 / Fragment: BRIL-3X / Mutation: M29W,H124I,R128L
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mus musculus (house mouse), (gene. exp.) Escherichia coli (E. coli)
Gene: Best1, Bmd1, Vmd2, cybC / Cell line (production host): HEK293 / Production host: Homo sapiens (human) / References: UniProt: O88870, UniProt: P0ABE7
#2: Chemical
ChemComp-CA / CALCIUM ION


Mass: 40.078 Da / Num. of mol.: 5 / Source method: obtained synthetically / Formula: Ca / Feature type: SUBJECT OF INVESTIGATION
#3: Chemical
ChemComp-CL / CHLORIDE ION


Mass: 35.453 Da / Num. of mol.: 10 / Source method: obtained synthetically / Formula: Cl
Has ligand of interestY
Has protein modificationN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Docked Pentameric structure of mouse bestrophin-1 in a partial open state
Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
21Mus musculus (house mouse)10090
31Escherichia coli (E. coli)562
Source (recombinant)Organism: Homo sapiens (human)
Buffer solutionpH: 8
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

MicroscopyModel: TFS GLACIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2750 nm / Nominal defocus min: 1000 nm
Image recordingElectron dose: 40 e/Å2 / Film or detector model: FEI FALCON IV (4k x 4k)

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Processing

EM softwareName: PHENIX / Version: 1.21rc1_5156: / Category: model refinement
CTF correctionType: NONE
3D reconstructionResolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 97720 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00315430
ELECTRON MICROSCOPYf_angle_d0.521010
ELECTRON MICROSCOPYf_dihedral_angle_d4.2672030
ELECTRON MICROSCOPYf_chiral_restr0.0372310
ELECTRON MICROSCOPYf_plane_restr0.0042615

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