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- PDB-9ayk: Cryo-EM structure of human Cav3.2 with ML218 -

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Basic information

Entry
Database: PDB / ID: 9ayk
TitleCryo-EM structure of human Cav3.2 with ML218
ComponentsVoltage-dependent T-type calcium channel subunit alpha-1H
KeywordsTRANSPORT PROTEIN / Cav3.2 / voltage gated calcium channel / cryo-EM / ML218
Function / homology
Function and homology information


low voltage-gated calcium channel activity / aldosterone biosynthetic process / cortisol biosynthetic process / positive regulation of acrosome reaction / voltage-gated monoatomic ion channel activity / NCAM1 interactions / myoblast fusion / calcium ion import / cellular response to potassium ion / : ...low voltage-gated calcium channel activity / aldosterone biosynthetic process / cortisol biosynthetic process / positive regulation of acrosome reaction / voltage-gated monoatomic ion channel activity / NCAM1 interactions / myoblast fusion / calcium ion import / cellular response to potassium ion / : / voltage-gated calcium channel complex / Mechanical load activates signaling by PIEZO1 and integrins in osteocytes / muscle organ development / regulation of heart contraction / calcium ion import across plasma membrane / Smooth Muscle Contraction / voltage-gated calcium channel activity / cellular response to hormone stimulus / muscle contraction / regulation of membrane potential / scaffold protein binding / metal ion binding / membrane / plasma membrane
Similarity search - Function
Voltage-dependent calcium channel, T-type, alpha-1 subunit / : / Voltage-dependent channel domain superfamily / Ion transport domain / Ion transport protein
Similarity search - Domain/homology
: / : / 1-O-OCTADECYL-SN-GLYCERO-3-PHOSPHOCHOLINE / CHOLESTEROL HEMISUCCINATE / Voltage-dependent T-type calcium channel subunit alpha-1H
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3 Å
AuthorsFan, X. / Huang, J. / Yan, N.
Funding support France, 1items
OrganizationGrant numberCountry
Human Frontier Science Program (HFSP)LT000754/2020-L France
CitationJournal: Cell Res / Year: 2024
Title: Structural basis for human Ca3.2 inhibition by selective antagonists.
Authors: Jian Huang / Xiao Fan / Xueqin Jin / Chen Lyu / Qinmeng Guo / Tao Liu / Jiaofeng Chen / Amaël Davakan / Philippe Lory / Nieng Yan /
Abstract: The Ca3.2 subtype of T-type calcium channels has been targeted for developing analgesics and anti-epileptics for its role in pain and epilepsy. Here we present the cryo-EM structures of Ca3.2 alone ...The Ca3.2 subtype of T-type calcium channels has been targeted for developing analgesics and anti-epileptics for its role in pain and epilepsy. Here we present the cryo-EM structures of Ca3.2 alone and in complex with four T-type calcium channel selective antagonists with overall resolutions ranging from 2.8 Å to 3.2 Å. The four compounds display two binding poses. ACT-709478 and TTA-A2 both place their cyclopropylphenyl-containing ends in the central cavity to directly obstruct ion flow, meanwhile extending their polar tails into the IV-I fenestration. TTA-P2 and ML218 project their 3,5-dichlorobenzamide groups into the II-III fenestration and place their hydrophobic tails in the cavity to impede ion permeation. The fenestration-penetrating mode immediately affords an explanation for the state-dependent activities of these antagonists. Structure-guided mutational analysis identifies several key residues that determine the T-type preference of these drugs. The structures also suggest the role of an endogenous lipid in stabilizing drug binding in the central cavity.
History
DepositionMar 8, 2024Deposition site: RCSB / Processing site: RCSB
Revision 1.0Apr 24, 2024Provider: repository / Type: Initial release
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Revision 1.0Apr 24, 2024Data content type: FSC / Data content type: FSC / Provider: repository / Type: Initial release
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Revision 1.1Jun 12, 2024Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first / _citation.page_last
Revision 1.2Oct 30, 2024Group: Data collection / Structure summary
Category: em_admin / pdbx_entry_details / pdbx_modification_feature
Item: _em_admin.last_update / _pdbx_entry_details.has_protein_modification
Revision 1.3May 14, 2025Group: Data collection / Category: em_admin / em_software / Item: _em_admin.last_update / _em_software.name
Revision 1.1May 14, 2025Data content type: EM metadata / Data content type: EM metadata / EM metadata / Group: Data processing / Experimental summary / Data content type: EM metadata / EM metadata / Category: em_admin / em_software / Data content type: EM metadata / EM metadata / Item: _em_admin.last_update / _em_software.name

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Voltage-dependent T-type calcium channel subunit alpha-1H
hetero molecules


Theoretical massNumber of molelcules
Total (without water)240,81914
Polymers234,8541
Non-polymers5,96513
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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Protein / Sugars , 2 types, 3 molecules A

#1: Protein Voltage-dependent T-type calcium channel subunit alpha-1H / Low-voltage-activated calcium channel alpha1 3.2 subunit / Voltage-gated calcium channel subunit alpha Cav3.2


Mass: 234853.609 Da / Num. of mol.: 1 / Fragment: UNP residues 1-491,772-2353
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CACNA1H / Production host: Homo sapiens (human) / References: UniProt: O95180
#3: 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.: 2 / 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 , 5 types, 11 molecules

#2: Chemical ChemComp-A1AHJ / 3,5-dichloro-N-{[(1R,5S,6r)-3-(3,3-dimethylbutyl)-3-azabicyclo[3.1.0]hexan-6-yl]methyl}benzamide


Mass: 369.329 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C19H26Cl2N2O / Feature type: SUBJECT OF INVESTIGATION
#4: Chemical ChemComp-CA / CALCIUM ION


Mass: 40.078 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Ca
#5: Chemical ChemComp-LPE / 1-O-OCTADECYL-SN-GLYCERO-3-PHOSPHOCHOLINE / LPC-ETHER


Mass: 510.708 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C26H57NO6P
#6: Chemical
ChemComp-Y01 / CHOLESTEROL HEMISUCCINATE


Mass: 486.726 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C31H50O4
#7: Chemical
ChemComp-JL3 / [(2~{R})-3-[2-azanylethoxy(oxidanyl)phosphoryl]oxy-2-pentadecanoyloxy-propyl] pentadecanoate / 1,2-Dipentadecanoyl-sn-glycero-3-phosphoethanolamine / PE(15:0/15:0)


Mass: 663.906 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C35H70NO8P

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Details

Has ligand of interestY
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

ComponentName: Voltage-dependent T-type calcium channel subunit alpha-1H
Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Homo sapiens (human)
Buffer solutionpH: 7.4
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
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: 1500 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 / Category: model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 126670 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0029048
ELECTRON MICROSCOPYf_angle_d0.43312252
ELECTRON MICROSCOPYf_dihedral_angle_d11.0631408
ELECTRON MICROSCOPYf_chiral_restr0.0341424
ELECTRON MICROSCOPYf_plane_restr0.0051475

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