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- PDB-9c49: Cryo-EM structure of Danio rerio voltage-sensing phosphatase (VSP... -

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Basic information

Entry
Database: PDB / ID: 9c49
TitleCryo-EM structure of Danio rerio voltage-sensing phosphatase (VSP) phosphatase domain
ComponentsPhosphatidylinositol-3,4,5-trisphosphate 3-phosphatase TPTE2
KeywordsMEMBRANE PROTEIN / phosphatase / voltage-sensing
Function / homology
Function and homology information


phosphatidylinositol-3,4,5-trisphosphate 3-phosphatase activity / phosphatidylinositol dephosphorylation / regulation of endocytosis / monoatomic ion channel activity / protein-tyrosine-phosphatase / cell projection / membrane / cytosol
Similarity search - Function
TPTE, protein tyrosine phosphatase-like catalytic domain / : / Tensin phosphatase, C2 domain / Tensin-type phosphatase domain / C2 domain of PTEN tumour-suppressor protein / Phosphatase tensin-type domain profile. / C2 tensin-type domain profile. / C2 domain of PTEN tumour-suppressor protein / Polymorphic toxin system, DSP-PTPase phosphatase / Voltage-dependent channel domain superfamily ...TPTE, protein tyrosine phosphatase-like catalytic domain / : / Tensin phosphatase, C2 domain / Tensin-type phosphatase domain / C2 domain of PTEN tumour-suppressor protein / Phosphatase tensin-type domain profile. / C2 tensin-type domain profile. / C2 domain of PTEN tumour-suppressor protein / Polymorphic toxin system, DSP-PTPase phosphatase / Voltage-dependent channel domain superfamily / Protein-tyrosine phosphatase, catalytic / Protein tyrosine phosphatase, catalytic domain motif / Tyrosine specific protein phosphatases active site. / Protein-tyrosine phosphatase, active site / Tyrosine-specific protein phosphatases domain / Tyrosine specific protein phosphatases domain profile. / Protein-tyrosine phosphatase-like / C2 domain superfamily / Ion transport domain / Ion transport protein
Similarity search - Domain/homology
Transmembrane phosphatase with tensin homology
Similarity search - Component
Biological speciesDanio rerio (zebrafish)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.97 Å
AuthorsZhang, L. / Brohawn, S.G.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM145869 United States
CitationJournal: Nat Commun / Year: 2024
Title: Coupling sensor to enzyme in the voltage sensing phosphatase.
Authors: Yawei Yu / Lin Zhang / Baobin Li / Zhu Fu / Stephen G Brohawn / Ehud Y Isacoff /
Abstract: Voltage-sensing phosphatases (VSPs) dephosphorylate phosphoinositide (PIP) signaling lipids in response to membrane depolarization. VSPs possess an S4-containing voltage sensor domain (VSD), ...Voltage-sensing phosphatases (VSPs) dephosphorylate phosphoinositide (PIP) signaling lipids in response to membrane depolarization. VSPs possess an S4-containing voltage sensor domain (VSD), resembling that of voltage-gated cation channels, and a lipid phosphatase domain (PD). The mechanism by which voltage turns on enzyme activity is unclear. Structural analysis and modeling suggest several sites of VSD-PD interaction that could couple voltage sensing to catalysis. Voltage clamp fluorometry reveals voltage-driven rearrangements in three sites implicated earlier in enzyme activation-the VSD-PD linker, gating loop and R loop-as well as the N-terminal domain, which has not yet been explored. N-terminus mutations perturb both rearrangements in the other segments and enzyme activity. Our results provide a model for a dynamic assembly by which S4 controls the catalytic site.
History
DepositionJun 3, 2024Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jul 10, 2024Provider: repository / Type: Initial release
Revision 1.1Aug 14, 2024Group: Data collection / Database references / Category: citation / citation_author / em_admin
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _citation_author.identifier_ORCID / _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Phosphatidylinositol-3,4,5-trisphosphate 3-phosphatase TPTE2
B: Phosphatidylinositol-3,4,5-trisphosphate 3-phosphatase TPTE2


Theoretical massNumber of molelcules
Total (without water)119,2812
Polymers119,2812
Non-polymers00
Water00
1


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

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Components

#1: Protein Phosphatidylinositol-3,4,5-trisphosphate 3-phosphatase TPTE2 / Transmembrane phosphatase with tensin homology


Mass: 59640.520 Da / Num. of mol.: 2 / Mutation: C302S
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Danio rerio (zebrafish) / Gene: tpte, tpip, vsp, wu:fd20e11, wu:fi24b06 / Production host: Komagataella pastoris (fungus) / References: UniProt: Q4V9E4, protein-tyrosine-phosphatase

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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-sensing phosphatase (Phosphatidylinositol-3,4,5-trisphosphate 3-phosphatase TPTE2) in lipid nanodisc
Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightValue: 0.119 MDa / Experimental value: NO
Source (natural)Organism: Danio rerio (zebrafish)
Source (recombinant)Organism: Komagataella pastoris (fungus)
Buffer solutionpH: 8
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMTRISC4H11NO31
2150 mMsodium chlorideNaCl1
35 mMethylene glycol-bis(beta-aminoethyl ether)-N,N,N,N-tetraacetic acidEGTA1
SpecimenConc.: 5.6 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Details: VSP reconstituted in MSP2N2 lipid nanodiscs with 2:1:1:0.2 DOPE:POPS:POPC:Brain PI(3,4)P2
Specimen supportGrid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: UltrAuFoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 4 K / Details: 3s blot, blot force 1

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Electron microscopy imaging

Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company
MicroscopyModel: FEI TALOS ARCTICA
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 1800 nm / Nominal defocus min: 600 nm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

EM software
IDNameCategory
2SerialEMimage acquisition
13cryoSPARC3D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 2.97 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 208690 / Symmetry type: POINT
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 49.17 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.01384880
ELECTRON MICROSCOPYf_angle_d1.33086592
ELECTRON MICROSCOPYf_chiral_restr0.0495700
ELECTRON MICROSCOPYf_plane_restr0.0067846
ELECTRON MICROSCOPYf_dihedral_angle_d5.0692648

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