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- PDB-8csl: Sub-tomogram averaging of erythrocyte ankyrin-1 complex -

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Basic information

Entry
Database: PDB / ID: 8csl
TitleSub-tomogram averaging of erythrocyte ankyrin-1 complex
Components
  • Ammonium transporter Rh type A
  • Ankyrin-1
  • Band 3 anion transport protein
  • Blood group Rh(CE) polypeptide
  • Glycophorin-A
  • Glycophorin-B
  • Protein 4.2
KeywordsTRANSPORT PROTEIN/STRUCTURAL PROTEIN / Membrane protein / ankyrin complex / TRANSPORT PROTEIN-STRUCTURAL PROTEIN complex
Function / homology
Function and homology information


methylammonium transmembrane transport / Defective RHAG causes regulator type Rh-null hemolytic anemia (RHN) / Rhesus blood group biosynthesis / methylammonium transmembrane transporter activity / Rhesus glycoproteins mediate ammonium transport / carbon dioxide transmembrane transport / carbon dioxide transmembrane transporter activity / ammonium homeostasis / spectrin-associated cytoskeleton / hemoglobin metabolic process ...methylammonium transmembrane transport / Defective RHAG causes regulator type Rh-null hemolytic anemia (RHN) / Rhesus blood group biosynthesis / methylammonium transmembrane transporter activity / Rhesus glycoproteins mediate ammonium transport / carbon dioxide transmembrane transport / carbon dioxide transmembrane transporter activity / ammonium homeostasis / spectrin-associated cytoskeleton / hemoglobin metabolic process / positive regulation of organelle organization / maintenance of epithelial cell apical/basal polarity / protein-glutamine gamma-glutamyltransferase activity / pH elevation / Defective SLC4A1 causes hereditary spherocytosis type 4 (HSP4), distal renal tubular acidosis (dRTA) and dRTA with hemolytic anemia (dRTA-HA) / negative regulation of urine volume / leak channel activity / NrCAM interactions / Neurofascin interactions / Bicarbonate transporters / ammonium transmembrane transport / intracellular monoatomic ion homeostasis / ankyrin-1 complex / ammonium channel activity / CHL1 interactions / monoatomic anion transmembrane transporter activity / plasma membrane phospholipid scrambling / cytoskeletal anchor activity / chloride:bicarbonate antiporter activity / solute:inorganic anion antiporter activity / bicarbonate transport / bicarbonate transmembrane transporter activity / monoatomic anion transport / M band / : / chloride transport / chloride transmembrane transporter activity / Interaction between L1 and Ankyrins / ankyrin binding / hemoglobin binding / spectrin binding / erythrocyte maturation / negative regulation of glycolytic process through fructose-6-phosphate / erythrocyte development / cortical cytoskeleton / exocytosis / axolemma / endoplasmic reticulum to Golgi vesicle-mediated transport / COPI-mediated anterograde transport / spleen development / protein-membrane adaptor activity / cytoskeleton organization / chloride transmembrane transport / sarcoplasmic reticulum / regulation of intracellular pH / Cell surface interactions at the vascular wall / protein localization to plasma membrane / carbon dioxide transport / Erythrocytes take up oxygen and release carbon dioxide / Erythrocytes take up carbon dioxide and release oxygen / sarcolemma / structural constituent of cytoskeleton / multicellular organismal-level iron ion homeostasis / transmembrane transport / cytoplasmic side of plasma membrane / Z disc / cell morphogenesis / blood coagulation / regulation of cell shape / virus receptor activity / ATPase binding / protein phosphatase binding / blood microparticle / basolateral plasma membrane / transmembrane transporter binding / postsynaptic membrane / cytoskeleton / neuron projection / structural molecule activity / enzyme binding / signal transduction / protein homodimerization activity / extracellular exosome / nucleoplasm / ATP binding / identical protein binding / membrane / plasma membrane / cytosol
Similarity search - Function
Blood group Rhesus C/E/D polypeptide / Glycophorin, conserved site / : / Glycophorin A / Glycophorin A signature. / Glycophorin / Ankyrin, UPA domain / UPA domain / : / Ammonium transporter AmtB-like domain ...Blood group Rhesus C/E/D polypeptide / Glycophorin, conserved site / : / Glycophorin A / Glycophorin A signature. / Glycophorin / Ankyrin, UPA domain / UPA domain / : / Ammonium transporter AmtB-like domain / Ammonium Transporter Family / Transglutaminase, N-terminal / Transglutaminase, C-terminal / Transglutaminase, active site / Protein-glutamine gamma-glutamyltransferase, animal / Transglutaminase, C-terminal domain superfamily / : / Transglutaminase family / Transglutaminase family, C-terminal ig like domain / Transglutaminases active site. / Domain present in ZO-1 and Unc5-like netrin receptors / ZU5 domain / ZU5 domain / ZU5 domain profile. / Transglutaminase-like superfamily / Transglutaminase/protease-like homologues / Transglutaminase-like / Anion exchange protein 1 / Ammonium/urea transporter / Transglutaminase-like superfamily / Anion exchange protein / Anion exchange, conserved site / Anion exchangers family signature 1. / Anion exchangers family signature 2. / Band 3 cytoplasmic domain / Band 3 cytoplasmic domain / Phosphotransferase/anion transporter / Bicarbonate transporter, eukaryotic / Bicarbonate transporter-like, transmembrane domain / HCO3- transporter integral membrane domain / Ankyrin repeat / Death domain profile. / DEATH domain, found in proteins involved in cell death (apoptosis). / Death domain / Death domain / Death-like domain superfamily / Ankyrin repeat / Papain-like cysteine peptidase superfamily / Ankyrin repeats (3 copies) / Ankyrin repeat profile. / Ankyrin repeat region circular profile. / ankyrin repeats / Ankyrin repeat / Ankyrin repeat-containing domain superfamily / Immunoglobulin E-set / Immunoglobulin-like fold
Similarity search - Domain/homology
Glycophorin-A / Band 3 anion transport protein / Glycophorin-B / Ankyrin-1 / Protein 4.2 / Blood group Rh(CE) polypeptide / Ammonium transporter Rh type A
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / subtomogram averaging / cryo EM / Resolution: 25 Å
AuthorsVallese, F. / Kim, K. / Yen, L.Y. / Johnston, J.D. / Noble, A.J. / Cali, T. / Clarke, O.B.
Funding support1items
OrganizationGrant numberCountry
Not funded
CitationJournal: Nat Struct Mol Biol / Year: 2022
Title: Architecture of the human erythrocyte ankyrin-1 complex.
Authors: Francesca Vallese / Kookjoo Kim / Laura Y Yen / Jake D Johnston / Alex J Noble / Tito Calì / Oliver Biggs Clarke /
Abstract: The stability and shape of the erythrocyte membrane is provided by the ankyrin-1 complex, but how it tethers the spectrin-actin cytoskeleton to the lipid bilayer and the nature of its association ...The stability and shape of the erythrocyte membrane is provided by the ankyrin-1 complex, but how it tethers the spectrin-actin cytoskeleton to the lipid bilayer and the nature of its association with the band 3 anion exchanger and the Rhesus glycoproteins remains unknown. Here we present structures of ankyrin-1 complexes purified from human erythrocytes. We reveal the architecture of a core complex of ankyrin-1, the Rhesus proteins RhAG and RhCE, the band 3 anion exchanger, protein 4.2, glycophorin A and glycophorin B. The distinct T-shaped conformation of membrane-bound ankyrin-1 facilitates recognition of RhCE and, unexpectedly, the water channel aquaporin-1. Together, our results uncover the molecular details of ankyrin-1 association with the erythrocyte membrane, and illustrate the mechanism of ankyrin-mediated membrane protein clustering.
History
DepositionMay 12, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jul 20, 2022Provider: repository / Type: Initial release
Revision 1.1Jul 27, 2022Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_PubMed / _citation.title
Revision 1.2Feb 14, 2024Group: Data collection / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / em_3d_fitting_list / pdbx_initial_refinement_model
Item: _em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id ..._em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id / _em_3d_fitting_list.source_name / _em_3d_fitting_list.type

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Ankyrin-1
W: Band 3 anion transport protein
K: Blood group Rh(CE) polypeptide
L: Ammonium transporter Rh type A
Q: Ammonium transporter Rh type A
X: Protein 4.2
P: Glycophorin-B
R: Glycophorin-A
a: Glycophorin-A
V: Band 3 anion transport protein
e: Band 3 anion transport protein
S: Glycophorin-A
b: Glycophorin-A
Y: Band 3 anion transport protein
f: Band 3 anion transport protein
T: Glycophorin-A
c: Glycophorin-A
Z: Band 3 anion transport protein
g: Band 3 anion transport protein


Theoretical massNumber of molelcules
Total (without water)1,238,74519
Polymers1,238,74519
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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Protein , 7 types, 19 molecules AWVeYfZgKLQXPRaSbTc

#1: Protein Ankyrin-1 / ANK-1 / Ankyrin-R / Erythrocyte ankyrin


Mass: 206522.625 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: P16157
#2: Protein
Band 3 anion transport protein / Anion exchange protein 1 / AE 1 / Anion exchanger 1 / Solute carrier family 4 member 1


Mass: 101883.859 Da / Num. of mol.: 7 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: P02730
#3: Protein Blood group Rh(CE) polypeptide / Rh polypeptide 1 / RhPI / Rh30A / RhIXB / Rhesus C/E antigens


Mass: 45598.918 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: P18577
#4: Protein Ammonium transporter Rh type A / Erythrocyte membrane glycoprotein Rh50 / Erythrocyte plasma membrane 50 kDa glycoprotein / Rh50A / ...Erythrocyte membrane glycoprotein Rh50 / Erythrocyte plasma membrane 50 kDa glycoprotein / Rh50A / Rhesus blood group family type A glycoprotein / Rh family type A glycoprotein / Rh type A glycoprotein / Rhesus blood group-associated ammonia channel / Rhesus blood group-associated glycoprotein


Mass: 44229.629 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: Q02094
#5: Protein Protein 4.2 / P4.2 / Erythrocyte membrane protein band 4.2 / Erythrocyte protein 4.2


Mass: 77096.914 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: P16452
#6: Protein Glycophorin-B / PAS-3 / SS-active sialoglycoprotein / Sialoglycoprotein delta


Mass: 9789.674 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: P06028
#7: Protein
Glycophorin-A / MN sialoglycoprotein / PAS-2 / Sialoglycoprotein alpha


Mass: 16348.433 Da / Num. of mol.: 6 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / References: UniProt: P02724

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: subtomogram averaging

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Sample preparation

ComponentName: Sub-tomogram average of ankyrin-1 complexes from native erythrocyte vesicles
Type: COMPLEX / Entity ID: all / Source: NATURAL
Source (natural)Organism: Homo sapiens (human)
Buffer solutionpH: 7.4 / Details: 130 mM KCl, 10 mM HEPES, pH 7.4
SpecimenConc.: 5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Details: Ghost membranes (~100 mg) were pelleted (6,000 x g, 10 minutes), washed 5 times in 130 mM KCl, 10 mM HEPES, pH 7.4, and then homogenized with the same buffer. The sample was sonicated using ...Details: Ghost membranes (~100 mg) were pelleted (6,000 x g, 10 minutes), washed 5 times in 130 mM KCl, 10 mM HEPES, pH 7.4, and then homogenized with the same buffer. The sample was sonicated using microprobe (amplitude = 30, pulse on = 5 sec, rest time = 10 sec, total pulse on = 50 sec). Large fragments are removed by centrifugation (6,000 x g, 10 minutes) and the supernatant was extruded (Avanti Polar Lipid) 10 times using a 400 nm filter. After extrusion, vesicles were collected by ultra-centrifugation at 35,000 rpm for 30 minutes (S120-AT3, Sorvall). The pellet, which contains vesicles, was resuspended at a final concentration of 5 mg/mL. All steps were performed at room temperature to facilitate vesicle formation.
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 5000 nm / Nominal defocus min: 3000 nm / Cs: 0.01 mm
Specimen holderCryogen: NITROGEN
Image recordingElectron dose: 3.14 e/Å2 / Avg electron dose per subtomogram: 113 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)
EM imaging opticsEnergyfilter slit width: 20 eV

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Processing

EM software
IDNameCategory
2Leginonimage acquisition
4WarpCTF correction
CTF correctionDetails: Warp / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 25 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 1596 / Symmetry type: POINT
EM volume selectionDetails: Picked using crYOLO, trained using manual picks. / Num. of tomograms: 100 / Num. of volumes extracted: 60029
Atomic model buildingProtocol: RIGID BODY FIT
Details: Erythrocyte ankyrin-1 complex (Class1, 8CS9) was rigid fit as a single body to the map using fitmap in UCSF Chimera, after which waters, ligands and sidechains were removed from the model.
Atomic model buildingPDB-ID: 8CS9

8cs9


Accession code: 8CS9 / Source name: PDB / Type: experimental model

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