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- EMDB-26145: Cryo-EM structure of human band 3-protein 4.2 complex in a loosel... -

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Basic information

Entry
Database: EMDB / ID: EMD-26145
TitleCryo-EM structure of human band 3-protein 4.2 complex in a loosely-bound conformation
Map databand 3-protein 4.2 complex in loosely-bound conformation
Sample
  • Complex: High-salt fraction 1 from human red blood cell membrane
KeywordsRed blood cell / Ankyrin complex / membrane protein / band 3 / protein 4.2
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.1 Å
AuthorsXia X / Liu SH / Zhou ZH
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Heart, Lung, and Blood Institute (NIH/NHLBI)R01GM071940 United States
CitationJournal: Nat Struct Mol Biol / Year: 2022
Title: Structure, dynamics and assembly of the ankyrin complex on human red blood cell membrane.
Authors: Xian Xia / Shiheng Liu / Z Hong Zhou /
Abstract: The cytoskeleton of a red blood cell (RBC) is anchored to the cell membrane by the ankyrin complex. This complex is assembled during RBC genesis and comprises primarily band 3, protein 4.2 and ...The cytoskeleton of a red blood cell (RBC) is anchored to the cell membrane by the ankyrin complex. This complex is assembled during RBC genesis and comprises primarily band 3, protein 4.2 and ankyrin, whose mutations contribute to numerous human inherited diseases. High-resolution structures of the ankyrin complex have been long sought-after to understand its assembly and disease-causing mutations. Here, we analyzed native complexes on the human RBC membrane by stepwise fractionation. Cryo-electron microscopy structures of nine band-3-associated complexes reveal that protein 4.2 stabilizes the cytoplasmic domain of band 3 dimer. In turn, the superhelix-shaped ankyrin binds to this protein 4.2 via ankyrin repeats (ARs) 6-13 and to another band 3 dimer via ARs 17-20, bridging two band 3 dimers in the ankyrin complex. Integration of these structures with both prior data and our biochemical data supports a model of ankyrin complex assembly during erythropoiesis and identifies interactions essential for the mechanical stability of RBC.
History
DepositionFeb 6, 2022-
Header (metadata) releaseJun 15, 2022-
Map releaseJun 15, 2022-
UpdateJan 17, 2024-
Current statusJan 17, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_26145.png

Downloads & links

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Map

FileDownload / File: emd_26145.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationband 3-protein 4.2 complex in loosely-bound conformation
Voxel sizeX=Y=Z: 1.1 Å
Density
Contour LevelBy AUTHOR: 0.01
Minimum - Maximum-0.027682312 - 0.07548089
Average (Standard dev.)0.00023032457 (±0.0023555094)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 352.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : High-salt fraction 1 from human red blood cell membrane

EntireName: High-salt fraction 1 from human red blood cell membrane
Components
  • Complex: High-salt fraction 1 from human red blood cell membrane

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Supramolecule #1: High-salt fraction 1 from human red blood cell membrane

SupramoleculeName: High-salt fraction 1 from human red blood cell membrane
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 280 KDa

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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

Concentration2.5 mg/mL
BufferpH: 7.5
Component:
ConcentrationFormulaName
10.0 mMNH2C(CH2OH)3HClTris-HClTris
300.0 mMNaClSodium chlorideSodium Chloride
1.0 mMHSCH2CH(OH)CH(OH)CH2SHDTT
0.015 %C24H46O11DDM
GridModel: UltrAuFoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: GOLD / Support film - topology: HOLEY
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 3.5 µm / Nominal defocus min: 1.8 µm / Nominal magnification: 81000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
TemperatureMin: 100.0 K / Max: 100.0 K
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number grids imaged: 1 / Number real images: 20842 / Average exposure time: 2.0 sec. / Average electron dose: 50.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 20740000
Startup modelType of model: NONE / Details: Generated from CryoSPARC
Initial angle assignmentType: OTHER / Software - Name: RELION (ver. 3.1)
Final angle assignmentType: OTHER / Software - Name: RELION (ver. 3.1)
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 4.1 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Number images used: 446313

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: AB INITIO MODEL / Overall B value: 192

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