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- PDB-8eog: Structure of the human L-type voltage-gated calcium channel Cav1.... -

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Basic information

Entry
Database: PDB / ID: 8eog
TitleStructure of the human L-type voltage-gated calcium channel Cav1.2 complexed with L-leucine
Components
  • (Voltage-dependent ...) x 2
  • Isoform 20 of Voltage-dependent L-type calcium channel subunit alpha-1C
KeywordsMEMBRANE PROTEIN / voltage-gated calcium channel / CaV alpha2delta / drug binding / gabapentinoid
Function / homology
Function and homology information


positive regulation of high voltage-gated calcium channel activity / calcium ion transmembrane transport via high voltage-gated calcium channel / L-type voltage-gated calcium channel complex / calcium ion import across plasma membrane / calcium channel regulator activity / voltage-gated calcium channel activity / T-tubule / metal ion binding / cytoplasm
Similarity search - Function
Voltage-dependent calcium channel, L-type, beta-3 subunit / Voltage-dependent L-type calcium channel subunit beta-3, SH3 domain / Voltage-dependent L-type calcium channel subunit beta-1-4, N-terminal A domain / Voltage-dependent calcium channel, L-type, beta subunit / Voltage gated calcium channel subunit beta domain 4Aa N terminal / VWA N-terminal / Voltage-dependent calcium channel, alpha-2/delta subunit, conserved region / VWA N-terminal / Neuronal voltage-dependent calcium channel alpha 2acd / Guanylate kinase/L-type calcium channel beta subunit ...Voltage-dependent calcium channel, L-type, beta-3 subunit / Voltage-dependent L-type calcium channel subunit beta-3, SH3 domain / Voltage-dependent L-type calcium channel subunit beta-1-4, N-terminal A domain / Voltage-dependent calcium channel, L-type, beta subunit / Voltage gated calcium channel subunit beta domain 4Aa N terminal / VWA N-terminal / Voltage-dependent calcium channel, alpha-2/delta subunit, conserved region / VWA N-terminal / Neuronal voltage-dependent calcium channel alpha 2acd / Guanylate kinase/L-type calcium channel beta subunit / Guanylate kinase / Guanylate kinase homologues. / von Willebrand factor type A domain / von Willebrand factor (vWF) type A domain / VWFA domain profile. / von Willebrand factor, type A / von Willebrand factor A-like domain superfamily / SH3-like domain superfamily / Src homology 3 (SH3) domain profile. / SH3 domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
CHOLESTEROL / LEUCINE / Chem-WNZ / Chem-WO9 / Voltage-dependent calcium channel subunit alpha-2/delta-1 / Voltage-dependent L-type calcium channel subunit beta-3 / Isoform 20 of Voltage-dependent L-type calcium channel subunit alpha-1C
Similarity search - Component
Biological speciesOryctolagus cuniculus (rabbit)
Homo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsChen, Z. / Mondal, A. / Abderemane-Ali, F. / Minor, D.L.
Funding support United States, 2items
OrganizationGrant numberCountry
National Institutes of Health/National Heart, Lung, and Blood Institute (NIH/NHLBI)HL080050 United States
National Institutes of Health/National Institute on Deafness and Other Communication Disorders (NIH/NIDCD)DC007664 United States
CitationJournal: Nature / Year: 2023
Title: EMC chaperone-Ca structure reveals an ion channel assembly intermediate.
Authors: Zhou Chen / Abhisek Mondal / Fayal Abderemane-Ali / Seil Jang / Sangeeta Niranjan / José L Montaño / Balyn W Zaro / Daniel L Minor /
Abstract: Voltage-gated ion channels (VGICs) comprise multiple structural units, the assembly of which is required for function. Structural understanding of how VGIC subunits assemble and whether chaperone ...Voltage-gated ion channels (VGICs) comprise multiple structural units, the assembly of which is required for function. Structural understanding of how VGIC subunits assemble and whether chaperone proteins are required is lacking. High-voltage-activated calcium channels (Cas) are paradigmatic multisubunit VGICs whose function and trafficking are powerfully shaped by interactions between pore-forming Ca1 or Ca2 Caα (ref. ), and the auxiliary Caβ and Caαδ subunits. Here we present cryo-electron microscopy structures of human brain and cardiac Ca1.2 bound with Caβ to a chaperone-the endoplasmic reticulum membrane protein complex (EMC)-and of the assembled Ca1.2-Caβ-Caαδ-1 channel. These structures provide a view of an EMC-client complex and define EMC sites-the transmembrane (TM) and cytoplasmic (Cyto) docks; interaction between these sites and the client channel causes partial extraction of a pore subunit and splays open the Caαδ-interaction site. The structures identify the Caαδ-binding site for gabapentinoid anti-pain and anti-anxiety drugs, show that EMC and Caαδ interactions with the channel are mutually exclusive, and indicate that EMC-to-Caαδ hand-off involves a divalent ion-dependent step and Ca1.2 element ordering. Disruption of the EMC-Ca complex compromises Ca function, suggesting that the EMC functions as a channel holdase that facilitates channel assembly. Together, the structures reveal a Ca assembly intermediate and EMC client-binding sites that could have wide-ranging implications for the biogenesis of VGICs and other membrane proteins.
History
DepositionOct 3, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0May 24, 2023Provider: repository / Type: Initial release
Revision 1.1Jul 26, 2023Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation_author.identifier_ORCID / _citation_author.name

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
D: Voltage-dependent calcium channel subunit alpha-2/delta-1
K: Isoform 20 of Voltage-dependent L-type calcium channel subunit alpha-1C
C: Voltage-dependent L-type calcium channel subunit beta-3
hetero molecules


Theoretical massNumber of molelcules
Total (without water)316,63021
Polymers311,3843
Non-polymers5,24618
Water543
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: mass spectrometry
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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Voltage-dependent ... , 2 types, 2 molecules DC

#1: Protein Voltage-dependent calcium channel subunit alpha-2/delta-1 / Voltage-gated calcium channel / Voltage-gated calcium channel subunit alpha-2/delta-1


Mass: 118941.695 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Oryctolagus cuniculus (rabbit) / Gene: CACNA2D1, CACNL2A, CCHL2A / Production host: Homo sapiens (human) / References: UniProt: P13806
#3: Protein Voltage-dependent L-type calcium channel subunit beta-3 / CAB3 / Calcium channel voltage-dependent subunit beta 3


Mass: 21667.154 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Oryctolagus cuniculus (rabbit) / Gene: CACNB3, CACNLB3 / Production host: Homo sapiens (human) / References: UniProt: P54286

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Protein , 1 types, 1 molecules K

#2: Protein Isoform 20 of Voltage-dependent L-type calcium channel subunit alpha-1C / Calcium channel / L type / alpha-1 polypeptide / isoform 1 / cardiac muscle / Voltage-gated calcium ...Calcium channel / L type / alpha-1 polypeptide / isoform 1 / cardiac muscle / Voltage-gated calcium channel subunit alpha Cav1.2


Mass: 170774.938 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CACNA1C, CACH2, CACN2, CACNL1A1, CCHL1A1 / Production host: Homo sapiens (human) / References: UniProt: Q13936-20

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Sugars , 3 types, 8 molecules

#4: Polysaccharide 2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2- ...2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 627.594 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DGlcpNAcb1-4DGlcpNAcb1-4DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/1,3,2/[a2122h-1b_1-5_2*NCC/3=O]/1-1-1/a4-b1_b4-c1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{}}}LINUCSPDB-CARE
#5: Polysaccharide 2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 424.401 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DGlcpNAcb1-4DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/1,2,1/[a2122h-1b_1-5_2*NCC/3=O]/1-1/a4-b1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{}}LINUCSPDB-CARE
#6: Sugar
ChemComp-NAG / 2-acetamido-2-deoxy-beta-D-glucopyranose / N-acetyl-beta-D-glucosamine / 2-acetamido-2-deoxy-beta-D-glucose / 2-acetamido-2-deoxy-D-glucose / 2-acetamido-2-deoxy-glucose / N-ACETYL-D-GLUCOSAMINE / N-Acetylglucosamine


Type: D-saccharide, beta linking / Mass: 221.208 Da / Num. of mol.: 5 / Source method: obtained synthetically / Formula: C8H15NO6
IdentifierTypeProgram
DGlcpNAcbCONDENSED IUPAC CARBOHYDRATE SYMBOLGMML 1.0
N-acetyl-b-D-glucopyranosamineCOMMON NAMEGMML 1.0
b-D-GlcpNAcIUPAC CARBOHYDRATE SYMBOLPDB-CARE 1.0
GlcNAcSNFG CARBOHYDRATE SYMBOLGMML 1.0

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Non-polymers , 7 types, 13 molecules

#7: Chemical ChemComp-CA / CALCIUM ION


Mass: 40.078 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: Ca
#8: Chemical ChemComp-NA / SODIUM ION


Mass: 22.990 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Formula: Na
#9: Chemical ChemComp-LEU / LEUCINE / Leucine


Type: L-peptide linking / Mass: 131.173 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C6H13NO2
#10: Chemical ChemComp-CLR / CHOLESTEROL / Cholesterol


Mass: 386.654 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C27H46O
#11: Chemical ChemComp-WNZ / (2R)-3-{[(R)-(2-aminoethoxy)(hydroxy)phosphoryl]oxy}-2-(dodecanoyloxy)propyl dodecanoate


Mass: 579.746 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C29H58NO8P
#12: Chemical ChemComp-WO9 / (2R)-3-{[(R)-(2-aminoethoxy)(hydroxy)phosphoryl]oxy}-2-(dodecanoyloxy)propyl heptadecanoate


Mass: 649.879 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C34H68NO8P
#13: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 3 / Source method: isolated from a natural source / Formula: H2O

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Details

Has ligand of interestN

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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

Component
IDNameTypeEntity IDParent-IDSource
1Ternary complex of human CaV alpha1C with rabbit CaV alpha2delta-1 and rabbit CaV beta3CELL#1-#30MULTIPLE SOURCES
2Human CaV alpha1CCOMPLEX#21RECOMBINANT
3Rabbit CaV alpha2delta-1COMPLEX#11RECOMBINANT
4Rabbit CaV beta3COMPLEX#31RECOMBINANT
Molecular weight
IDEntity assembly-IDValue (°)Experimental value
11
220.187 MDaYES
330.125 MDaYES
440.1 MDaYES
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
21Homo sapiens (human)9606
32Homo sapiens (human)9606
43Oryctolagus cuniculus (rabbit)9986
54Oryctolagus cuniculus (rabbit)9986
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-ID
21Homo sapiens (human)9606
32Homo sapiens (human)9606
43Homo sapiens (human)9606
54Homo sapiens (human)9606
Buffer solutionpH: 8
SpecimenConc.: 2.7 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K

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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 FIELDBright-field microscopy / Nominal magnification: 105000 X / Nominal defocus max: 1700 nm / Nominal defocus min: 900 nm / Cs: 2.7 mm
Image recordingElectron dose: 46 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.20.1_4487: / Classification: refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 269802 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00220159
ELECTRON MICROSCOPYf_angle_d0.56727333
ELECTRON MICROSCOPYf_dihedral_angle_d17.3687397
ELECTRON MICROSCOPYf_chiral_restr0.0423125
ELECTRON MICROSCOPYf_plane_restr0.0043431

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