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- PDB-7t8t: CryoEM structure of PLCg1 -

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Basic information

Entry
Database: PDB / ID: 7t8t
TitleCryoEM structure of PLCg1
Components1-phosphatidylinositol 4,5-bisphosphate phosphodiesterase gamma
KeywordsHYDROLASE / 1-phosphatidylinositol 4 / 5-bisphosphate phosphodiesterase gamma-1 cryo EM
Function / homology
Function and homology information


phosphoinositide phospholipase C / phospholipid catabolic process / phosphatidylinositol phospholipase C activity / intracellular signal transduction / calcium ion binding
Similarity search - Function
1-phosphatidylinositol 4,5-bisphosphate phosphodiesterase gamma-1, SH3 domain / Phosphatidylinositol-4, 5-bisphosphate phosphodiesterase gamma / PLC-gamma, C-terminal SH2 domain / PLC-gamma, N-terminal SH2 domain / Phosphoinositide phospholipase C family / Phospholipase C, phosphatidylinositol-specific, Y domain / Phosphatidylinositol-specific phospholipase C, Y domain / Phosphatidylinositol-specific phospholipase Y-box domain profile. / Phospholipase C, catalytic domain (part); domain Y / Phosphatidylinositol-specific phospholipase C, X domain ...1-phosphatidylinositol 4,5-bisphosphate phosphodiesterase gamma-1, SH3 domain / Phosphatidylinositol-4, 5-bisphosphate phosphodiesterase gamma / PLC-gamma, C-terminal SH2 domain / PLC-gamma, N-terminal SH2 domain / Phosphoinositide phospholipase C family / Phospholipase C, phosphatidylinositol-specific, Y domain / Phosphatidylinositol-specific phospholipase C, Y domain / Phosphatidylinositol-specific phospholipase Y-box domain profile. / Phospholipase C, catalytic domain (part); domain Y / Phosphatidylinositol-specific phospholipase C, X domain / Phosphatidylinositol-specific phospholipase C, X domain / Phospholipase C, catalytic domain (part); domain X / Phosphatidylinositol-specific phospholipase X-box domain profile. / PLC-like phosphodiesterase, TIM beta/alpha-barrel domain superfamily / Protein kinase C conserved region 2 (CalB) / C2 domain / C2 domain / C2 domain profile. / PH domain / C2 domain superfamily / PH domain profile. / Pleckstrin homology domain. / Pleckstrin homology domain / SH3 domain / SH2 domain / Src homology 2 (SH2) domain profile. / Src homology 2 domains / SH2 domain / Src homology 3 domains / EF-Hand 1, calcium-binding site / SH2 domain superfamily / EF-hand calcium-binding domain. / EF-hand calcium-binding domain profile. / SH3-like domain superfamily / Src homology 3 (SH3) domain profile. / SH3 domain / EF-hand domain / EF-hand domain pair / PH-like domain superfamily
Similarity search - Domain/homology
1-phosphatidylinositol 4,5-bisphosphate phosphodiesterase gamma
Similarity search - Component
Biological speciesRattus norvegicus (Norway rat)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.68 Å
AuthorsEndo-Streeter, S. / Sondek, J.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01-GM057391 United States
CitationJournal: To Be Published
Title: CryoEM structure of PLCg1
Authors: Endo-Streeter, S. / Sondek, J.
History
DepositionDec 17, 2021Deposition site: RCSB / Processing site: RCSB
Revision 1.0Dec 21, 2022Provider: repository / Type: Initial release
Revision 1.1Jun 5, 2024Group: Data collection / Refinement description
Category: chem_comp_atom / chem_comp_bond / em_3d_fitting_list
Item: _em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id ..._em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id / _em_3d_fitting_list.source_name / _em_3d_fitting_list.type

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: 1-phosphatidylinositol 4,5-bisphosphate phosphodiesterase gamma
hetero molecules


Theoretical massNumber of molelcules
Total (without water)138,2252
Polymers138,1851
Non-polymers401
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein 1-phosphatidylinositol 4,5-bisphosphate phosphodiesterase gamma


Mass: 138185.391 Da / Num. of mol.: 1 / Mutation: H335A
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Rattus norvegicus (Norway rat) / Gene: Plcg1, rCG_32419 / Production host: Trichoplusia ni (cabbage looper)
References: UniProt: G3V845, phosphoinositide phospholipase C
#2: Chemical ChemComp-CA / CALCIUM ION


Mass: 40.078 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Ca / Feature type: SUBJECT OF INVESTIGATION
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: 1-phosphatidylinositol 4,5-bisphosphate phosphodiesterase gamma-1
Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Rattus norvegicus (Norway rat)
Source (recombinant)Organism: Trichoplusia ni (cabbage looper)
Buffer solutionpH: 7.4
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMHEPESC8H18N2O4S1
2100 mMNaClNaCl1
310 mMMgCl2MgCl21
42 mMDTTC4H10O2S21
50.15 mMn-Dodecyl-B-D-MaltosideC24H46O111
SpecimenConc.: 10 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: C-flat-1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE-PROPANE / Humidity: 95 % / Chamber temperature: 295 K

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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 / Cs: 2.7 mm
Specimen holderCryogen: NITROGEN
Image recordingElectron dose: 55 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 4450
Image scansWidth: 5760 / Height: 4092

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Processing

Software
NameVersionClassificationNB
phenix.real_space_refine1.19.2_4158refinement
PHENIX1.19.2_4158refinement
EM software
IDNameVersionCategoryDetails
1cryoSPARC3.2particle selection
7UCSF Chimera1.15model fittingWindows version
9PHENIX1.19.2_4158model refinement
13cryoSPARC3.23D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.68 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 238458 / Num. of class averages: 13 / Symmetry type: POINT
Atomic model buildingB value: 135.93 / Protocol: FLEXIBLE FIT / Space: REAL / Target criteria: CC fit and model metrics
Atomic model buildingPDB-ID: 6PBC

6pbc


Pdb chain-ID: A / Accession code: 6PBC / Pdb chain residue range: 21-1215 / Source name: PDB / Type: experimental model
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 135.93 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00219013
ELECTRON MICROSCOPYf_angle_d0.467812185
ELECTRON MICROSCOPYf_chiral_restr0.03911307
ELECTRON MICROSCOPYf_plane_restr0.00311584
ELECTRON MICROSCOPYf_dihedral_angle_d3.80271193

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