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- PDB-8eoi: Structure of a human EMC:human Cav1.2 channel complex in GDN detergent -

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Basic information

Entry
Database: PDB / ID: 8eoi
TitleStructure of a human EMC:human Cav1.2 channel complex in GDN detergent
Components
  • (ER membrane protein complex subunit ...) x 9
  • (Voltage-dependent L-type calcium channel subunit ...) x 2
KeywordsMEMBRANE PROTEIN / endoplasmic reticulum membrane protein complex / voltage-gated calcium channel / holdase / biogenesis
Function / homology
Function and homology information


extrinsic component of endoplasmic reticulum membrane / inorganic cation transmembrane transporter activity / EMC complex / omegasome membrane / protein insertion into ER membrane by stop-transfer membrane-anchor sequence / voltage-gated calcium channel activity involved in AV node cell action potential / voltage-gated calcium channel activity involved in cardiac muscle cell action potential / immune system development / positive regulation of high voltage-gated calcium channel activity / magnesium ion transport ...extrinsic component of endoplasmic reticulum membrane / inorganic cation transmembrane transporter activity / EMC complex / omegasome membrane / protein insertion into ER membrane by stop-transfer membrane-anchor sequence / voltage-gated calcium channel activity involved in AV node cell action potential / voltage-gated calcium channel activity involved in cardiac muscle cell action potential / immune system development / positive regulation of high voltage-gated calcium channel activity / magnesium ion transport / positive regulation of adenylate cyclase activity / membrane depolarization during atrial cardiac muscle cell action potential / Phase 2 - plateau phase / calcium ion transmembrane transport via high voltage-gated calcium channel / tail-anchored membrane protein insertion into ER membrane / Miscellaneous transport and binding events / cobalt ion transmembrane transporter activity / positive regulation of muscle contraction / ferrous iron transmembrane transporter activity / membrane depolarization during AV node cell action potential / high voltage-gated calcium channel activity / copper ion transport / cardiac conduction / L-type voltage-gated calcium channel complex / magnesium ion transmembrane transporter activity / membrane depolarization during cardiac muscle cell action potential / cell communication by electrical coupling involved in cardiac conduction / regulation of ventricular cardiac muscle cell action potential / NCAM1 interactions / camera-type eye development / cardiac muscle cell action potential involved in contraction / embryonic forelimb morphogenesis / calcium ion transport into cytosol / voltage-gated calcium channel complex / Phase 0 - rapid depolarisation / alpha-actinin binding / regulation of heart rate by cardiac conduction / calcium ion import across plasma membrane / autophagosome assembly / RHOA GTPase cycle / voltage-gated calcium channel activity / regulation of cardiac muscle contraction by regulation of the release of sequestered calcium ion / positive regulation of endothelial cell proliferation / calcium channel regulator activity / positive regulation of endothelial cell migration / Regulation of insulin secretion / postsynaptic density membrane / calcium ion transmembrane transport / Z disc / positive regulation of angiogenesis / Adrenaline,noradrenaline inhibits insulin secretion / heart development / positive regulation of cytosolic calcium ion concentration / carbohydrate binding / early endosome membrane / angiogenesis / perikaryon / early endosome / calmodulin binding / postsynaptic density / cilium / Golgi membrane / apoptotic process / dendrite / endoplasmic reticulum membrane / endoplasmic reticulum / Golgi apparatus / protein-containing complex / extracellular region / nucleoplasm / metal ion binding / membrane / plasma membrane / cytoplasm
Similarity search - Function
ER membrane protein complex subunit 8/9 / : / Uncharacterised protein family (UPF0172) / EMC2 TPR-like repeat domain / TMEM85/ER membrane protein complex subunit 4 / ER membrane protein complex subunit 4 / ER membrane protein complex subunit 7, beta-sandwich domain / ER membrane protein complex subunit 7 / ER membrane protein complex subunit 7, beta-sandwich domain / ER membrane protein complex subunit 6 ...ER membrane protein complex subunit 8/9 / : / Uncharacterised protein family (UPF0172) / EMC2 TPR-like repeat domain / TMEM85/ER membrane protein complex subunit 4 / ER membrane protein complex subunit 4 / ER membrane protein complex subunit 7, beta-sandwich domain / ER membrane protein complex subunit 7 / ER membrane protein complex subunit 7, beta-sandwich domain / ER membrane protein complex subunit 6 / ER membrane protein complex subunit 3 / ER membrane protein complex subunit 1, C-terminal / Membrane magnesium transporter / ER membrane protein complex subunit 1 / ER membrane protein complex subunit 6-like / EMC6 / ER membrane protein complex subunit 1, C-terminal / Membrane magnesium transporter / ER membrane protein complex subunit 10 / ER membrane protein complex subunit 2-like / Integral membrane protein EMC3/TMCO1-like / Integral membrane protein EMC3/TMCO1-like / Integral membrane protein DUF106 / Voltage-dependent calcium channel, L-type, beta-3 subunit / Voltage-dependent L-type calcium channel subunit beta-3, SH3 domain / Voltage-dependent calcium channel, L-type, alpha-1C subunit / Voltage-dependent calcium channel, L-type, beta subunit / Voltage-dependent L-type calcium channel subunit beta-1-4, N-terminal A domain / Voltage gated calcium channel subunit beta domain 4Aa N terminal / Voltage-gated calcium channel subunit alpha, C-terminal / Voltage-gated calcium channel subunit alpha, C-term / Voltage-dependent calcium channel, L-type, alpha-1 subunit / Voltage-gated potassium channels. Chain C / Voltage-dependent calcium channel, alpha-1 subunit, IQ domain / Voltage gated calcium channel IQ domain / Voltage gated calcium channel IQ domain / Voltage-dependent calcium channel, alpha-1 subunit / Carbohydrate-binding-like fold / Voltage-dependent L-type calcium channel, IQ-associated domain / Voltage-dependent L-type calcium channel, IQ-associated / : / Pyrrolo-quinoline quinone repeat / Outer membrane protein assembly factor BamB / Guanylate kinase/L-type calcium channel beta subunit / Guanylate kinase / Guanylate kinase homologues. / Quinoprotein alcohol dehydrogenase-like superfamily / SH3 Domains / Voltage-dependent channel domain superfamily / TPR repeat region circular profile. / TPR repeat profile. / MPN domain / MPN domain profile. / Tetratricopeptide repeats / Tetratricopeptide repeat / Four Helix Bundle (Hemerythrin (Met), subunit A) / SH3 type barrels. / SH3-like domain superfamily / Src homology 3 (SH3) domain profile. / SH3 domain / Ion transport domain / Ion transport protein / Tetratricopeptide-like helical domain superfamily / Roll / P-loop containing nucleotide triphosphate hydrolases / WD40/YVTN repeat-like-containing domain superfamily / Up-down Bundle / Rossmann fold / P-loop containing nucleoside triphosphate hydrolase / 3-Layer(aba) Sandwich / Mainly Beta / Mainly Alpha / Alpha Beta
Similarity search - Domain/homology
ER membrane protein complex subunit 8 / Voltage-dependent L-type calcium channel subunit beta-3 / Voltage-dependent L-type calcium channel subunit alpha-1C / ER membrane protein complex subunit 2 / ER membrane protein complex subunit 4 / ER membrane protein complex subunit 10 / ER membrane protein complex subunit 5 / ER membrane protein complex subunit 1 / ER membrane protein complex subunit 6 / Endoplasmic reticulum membrane protein complex subunit 7 / ER membrane protein complex subunit 3
Similarity search - Component
Biological speciesHomo sapiens (human)
Oryctolagus cuniculus (rabbit)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.4 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
Revision 1.2Nov 6, 2024Group: Data collection / Structure summary
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / em_admin / pdbx_entry_details / pdbx_modification_feature
Item: _em_admin.last_update / _pdbx_entry_details.has_protein_modification

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: ER membrane protein complex subunit 1
B: ER membrane protein complex subunit 2
C: ER membrane protein complex subunit 3
E: ER membrane protein complex subunit 5
F: ER membrane protein complex subunit 6
G: ER membrane protein complex subunit 7
H: ER membrane protein complex subunit 8
I: ER membrane protein complex subunit 10
K: Voltage-dependent L-type calcium channel subunit alpha-1C
J: Voltage-dependent L-type calcium channel subunit beta-3
D: ER membrane protein complex subunit 4
hetero molecules


Theoretical massNumber of molelcules
Total (without water)483,94516
Polymers481,90611
Non-polymers2,0395
Water00
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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ER membrane protein complex subunit ... , 9 types, 9 molecules ABCEFGHID

#1: Protein ER membrane protein complex subunit 1


Mass: 109598.438 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: EMC1, KIAA0090, PSEC0263 / Production host: Homo sapiens (human) / References: UniProt: Q8N766
#2: Protein ER membrane protein complex subunit 2 / Tetratricopeptide repeat protein 35 / TPR repeat protein 35


Mass: 34250.797 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: EMC2, KIAA0103, TTC35 / Production host: Homo sapiens (human) / References: UniProt: Q15006
#3: Protein ER membrane protein complex subunit 3 / Transmembrane protein 111


Mass: 29722.598 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: EMC3, TMEM111 / Production host: Homo sapiens (human) / References: UniProt: Q9P0I2
#4: Protein ER membrane protein complex subunit 5 / Membrane magnesium transporter 1 / Transmembrane protein 32


Mass: 11425.147 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: MMGT1, EMC5, TMEM32 / Production host: Homo sapiens (human) / References: UniProt: Q8N4V1
#5: Protein ER membrane protein complex subunit 6 / Transmembrane protein 93


Mass: 11014.019 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: EMC6, TMEM93 / Production host: Homo sapiens (human) / References: UniProt: Q9BV81
#6: Protein ER membrane protein complex subunit 7


Mass: 13151.044 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: EMC7, C11orf3, C15orf24, HT022, UNQ905/PRO1926 / Production host: Homo sapiens (human) / References: UniProt: Q9NPA0
#7: Protein ER membrane protein complex subunit 8 / Neighbor of COX4 / Protein FAM158B


Mass: 23578.764 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: EMC8, C16orf2, C16orf4, COX4AL, COX4NB, FAM158B, NOC4 / Production host: Homo sapiens (human) / References: UniProt: O43402
#8: Protein ER membrane protein complex subunit 10 / Hematopoietic signal peptide-containing membrane domain-containing protein 1


Mass: 17003.912 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: EMC10, C19orf63, INM02, UNQ764/PRO1556 / Production host: Homo sapiens (human) / References: UniProt: Q5UCC4
#11: Protein ER membrane protein complex subunit 4 / Cell proliferation-inducing gene 17 protein / Transmembrane protein 85


Mass: 18790.051 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: EMC4, TMEM85, HSPC184, PIG17 / Production host: Homo sapiens (human) / References: UniProt: Q5J8M3

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Voltage-dependent L-type calcium channel subunit ... , 2 types, 2 molecules KJ

#9: Protein 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: 176678.766 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
#10: Protein Voltage-dependent L-type calcium channel subunit beta-3 / CAB3 / Calcium channel voltage-dependent subunit beta 3


Mass: 36692.055 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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Sugars , 2 types, 4 molecules

#12: 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.: 3
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
#13: 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


Type: D-saccharide, beta linking / Mass: 221.208 Da / Num. of mol.: 1 / 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 , 1 types, 1 molecules

#14: Chemical ChemComp-9Z9 / (3beta,14beta,17beta,25R)-3-[4-methoxy-3-(methoxymethyl)butoxy]spirost-5-en


Mass: 544.805 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C34H56O5 / Comment: detergent*YM

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Details

Has ligand of interestN
Has protein modificationY

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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 the human ER membrane protein complex (EMC) with human CaV alpha1C and rabbit CaV beta3COMPLEX#1-#110MULTIPLE SOURCES
2Human CaV alpha1CCOMPLEX#91RECOMBINANT
3ER membrane protein complex subunit 1COMPLEX#11RECOMBINANT
4Rabbit CaV beta3COMPLEX#101RECOMBINANT
Molecular weight
IDEntity assembly-IDValue (°)Experimental value
11
220.187 MDaYES
330.112 MDaYES
440.1 MDaYES
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
21Homo sapiens (human)9606
32Homo sapiens (human)9606
43Homo sapiens (human)9606
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.: 1.7 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 FIELD / 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: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 487067 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00229381
ELECTRON MICROSCOPYf_angle_d0.47439827
ELECTRON MICROSCOPYf_dihedral_angle_d11.4410754
ELECTRON MICROSCOPYf_chiral_restr0.044513
ELECTRON MICROSCOPYf_plane_restr0.0045011

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