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- PDB-8ouo: Human TPC2 in Complex with Antagonist (S)-SG-094 -

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Basic information

Entry
Database: PDB / ID: 8ouo
TitleHuman TPC2 in Complex with Antagonist (S)-SG-094
ComponentsTwo pore channel protein 2
KeywordsMETAL TRANSPORT / TPC2 / two-pore channel / ion channel / inhibitor / homodimer
Function / homology
Function and homology information


endosome to lysosome transport of low-density lipoprotein particle / negative regulation of developmental pigmentation / intracellular pH reduction / intracellularly phosphatidylinositol-3,5-bisphosphate-gated monatomic cation channel activity / ligand-gated sodium channel activity / NAADP-sensitive calcium-release channel activity / melanosome membrane / regulation of exocytosis / response to vitamin D / endolysosome membrane ...endosome to lysosome transport of low-density lipoprotein particle / negative regulation of developmental pigmentation / intracellular pH reduction / intracellularly phosphatidylinositol-3,5-bisphosphate-gated monatomic cation channel activity / ligand-gated sodium channel activity / NAADP-sensitive calcium-release channel activity / melanosome membrane / regulation of exocytosis / response to vitamin D / endolysosome membrane / phosphatidylinositol-3,5-bisphosphate binding / monoatomic ion channel complex / lysosome organization / sodium ion transmembrane transport / smooth muscle contraction / voltage-gated calcium channel activity / release of sequestered calcium ion into cytosol / monoatomic ion transmembrane transport / regulation of autophagy / calcium-mediated signaling / calcium channel activity / endocytosis involved in viral entry into host cell / Stimuli-sensing channels / intracellular calcium ion homeostasis / late endosome membrane / receptor-mediated endocytosis of virus by host cell / lysosome / endosome membrane / lysosomal membrane / protein kinase binding / identical protein binding / cytosol
Similarity search - Function
Two pore channel protein 2 / Voltage-dependent channel domain superfamily / Ion transport domain / Ion transport protein
Similarity search - Domain/homology
1,2-DIACYL-SN-GLYCERO-3-PHOSHOCHOLINE / di-heneicosanoyl phosphatidyl choline / Chem-Q7G / : / CHOLESTEROL HEMISUCCINATE / Two pore channel protein 2
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3 Å
AuthorsChi, G. / Pike, A.C.W. / Maclean, E.M. / Li, H. / Mukhopadhyay, S.M.M. / Bohstedt, T. / Wang, D. / McKinley, G. / Fernandez-Cid, A. / Duerr, K.
Funding support United Kingdom, 1items
OrganizationGrant numberCountry
Wellcome Trust United Kingdom
CitationJournal: Structure / Year: 2024
Title: Structural basis for inhibition of the lysosomal two-pore channel TPC2 by a small molecule antagonist.
Authors: Gamma Chi / Dawid Jaślan / Veronika Kudrina / Julia Böck / Huanyu Li / Ashley C W Pike / Susanne Rautenberg / Einar Krogsaeter / Tina Bohstedt / Dong Wang / Gavin McKinley / Alejandra ...Authors: Gamma Chi / Dawid Jaślan / Veronika Kudrina / Julia Böck / Huanyu Li / Ashley C W Pike / Susanne Rautenberg / Einar Krogsaeter / Tina Bohstedt / Dong Wang / Gavin McKinley / Alejandra Fernandez-Cid / Shubhashish M M Mukhopadhyay / Nicola A Burgess-Brown / Marco Keller / Franz Bracher / Christian Grimm / Katharina L Dürr /
Abstract: Two pore channels are lysosomal cation channels with crucial roles in tumor angiogenesis and viral release from endosomes. Inhibition of the two-pore channel 2 (TPC2) has emerged as potential ...Two pore channels are lysosomal cation channels with crucial roles in tumor angiogenesis and viral release from endosomes. Inhibition of the two-pore channel 2 (TPC2) has emerged as potential therapeutic strategy for the treatment of cancers and viral infections, including Ebola and COVID-19. Here, we demonstrate that antagonist SG-094, a synthetic analog of the Chinese alkaloid medicine tetrandrine with increased potency and reduced toxicity, induces asymmetrical structural changes leading to a single binding pocket at only one intersubunit interface within the asymmetrical dimer. Supported by functional characterization of mutants by Ca imaging and patch clamp experiments, we identify key residues in S1 and S4 involved in compound binding to the voltage sensing domain II. SG-094 arrests IIS4 in a downward shifted state which prevents pore opening via the IIS4/S5 linker, hence resembling gating modifiers of canonical VGICs. These findings may guide the rational development of new therapeutics antagonizing TPC2 activity.
History
DepositionApr 24, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Jun 12, 2024Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Two pore channel protein 2
B: Two pore channel protein 2
hetero molecules


Theoretical massNumber of molelcules
Total (without water)180,65716
Polymers170,5712
Non-polymers10,08714
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area15480 Å2
ΔGint-131 kcal/mol
Surface area57790 Å2
MethodPISA

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Components

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Protein , 1 types, 2 molecules AB

#1: Protein Two pore channel protein 2 / Two pore calcium channel protein 2


Mass: 85285.430 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: TPCN2, TPC2 / Production host: Homo sapiens (human) / References: UniProt: Q8NHX9

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Non-polymers , 5 types, 14 molecules

#2: Chemical ChemComp-PLD / di-heneicosanoyl phosphatidyl choline


Mass: 875.313 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C50H101NO8P
#3: Chemical ChemComp-Q7G / 2-{[(4-O-alpha-D-glucopyranosyl-alpha-D-glucopyranosyl)oxy]methyl}-4-{[(3beta,9beta,14beta,17beta,25R)-spirost-5-en-3-yl]oxy}butyl 4-O-alpha-D-glucopyranosyl-alpha-D-glucopyranoside


Mass: 1165.315 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C56H92O25
#4: Chemical ChemComp-W4E / (1S)-6-methoxy-2-methyl-7-phenoxy-1-[(4-phenoxyphenyl)methyl]-3,4-dihydro-1H-isoquinoline / SG-094


Mass: 451.556 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C30H29NO3 / Feature type: SUBJECT OF INVESTIGATION
#5: Chemical ChemComp-PCF / 1,2-DIACYL-SN-GLYCERO-3-PHOSHOCHOLINE


Mass: 734.039 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C40H80NO8P
#6: Chemical
ChemComp-Y01 / CHOLESTEROL HEMISUCCINATE


Mass: 486.726 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: C31H50O4

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Details

Has ligand of interestY

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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: Homodimeric complex of human Two-pore channel 2 (TPC2)
Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Molecular weightValue: 0.15 MDa / Experimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Homo sapiens (human)
Buffer solutionpH: 7.5
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: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2400 nm / Nominal defocus min: 800 nm
Image recordingElectron dose: 35 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 109417 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00411045
ELECTRON MICROSCOPYf_angle_d0.50515072
ELECTRON MICROSCOPYf_dihedral_angle_d10.8364150
ELECTRON MICROSCOPYf_chiral_restr0.0391764
ELECTRON MICROSCOPYf_plane_restr0.0031778

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