+Open data
-Basic information
Entry | Database: PDB / ID: 6e1p | |||||||||
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Title | Structure of AtTPC1(DDE) in state 2 | |||||||||
Components | Two pore calcium channel protein 1 | |||||||||
Keywords | MEMBRANE PROTEIN / Two-pore channel | |||||||||
Function / homology | Function and homology information regulation of jasmonic acid biosynthetic process / seed germination / regulation of stomatal movement / plant-type vacuole / vacuole / vacuolar membrane / monoatomic ion channel complex / voltage-gated calcium channel activity / calcium-mediated signaling / calcium ion transport ...regulation of jasmonic acid biosynthetic process / seed germination / regulation of stomatal movement / plant-type vacuole / vacuole / vacuolar membrane / monoatomic ion channel complex / voltage-gated calcium channel activity / calcium-mediated signaling / calcium ion transport / calcium ion binding / Golgi apparatus / identical protein binding / plasma membrane / cytosol Similarity search - Function | |||||||||
Biological species | Arabidopsis thaliana (thale cress) | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.7 Å | |||||||||
Authors | Kintzer, A.F. / Green, E.M. / Cheng, Y. / Stroud, R.M. | |||||||||
Funding support | United States, 2items
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Citation | Journal: Proc Natl Acad Sci U S A / Year: 2018 Title: Structural basis for activation of voltage sensor domains in an ion channel TPC1. Authors: Alexander F Kintzer / Evan M Green / Pawel K Dominik / Michael Bridges / Jean-Paul Armache / Dawid Deneka / Sangwoo S Kim / Wayne Hubbell / Anthony A Kossiakoff / Yifan Cheng / Robert M Stroud / Abstract: Voltage-sensing domains (VSDs) couple changes in transmembrane electrical potential to conformational changes that regulate ion conductance through a central channel. Positively charged amino acids ...Voltage-sensing domains (VSDs) couple changes in transmembrane electrical potential to conformational changes that regulate ion conductance through a central channel. Positively charged amino acids inside each sensor cooperatively respond to changes in voltage. Our previous structure of a TPC1 channel captured an example of a resting-state VSD in an intact ion channel. To generate an activated-state VSD in the same channel we removed the luminal inhibitory Ca-binding site (Ca), which shifts voltage-dependent opening to more negative voltage and activation at 0 mV. Cryo-EM reveals two coexisting structures of the VSD, an intermediate state 1 that partially closes access to the cytoplasmic side but remains occluded on the luminal side and an intermediate activated state 2 in which the cytoplasmic solvent access to the gating charges closes, while luminal access partially opens. Activation can be thought of as moving a hydrophobic insulating region of the VSD from the external side to an alternate grouping on the internal side. This effectively moves the gating charges from the inside potential to that of the outside. Activation also requires binding of Ca to a cytoplasmic site (Ca). An X-ray structure with Ca removed and a near-atomic resolution cryo-EM structure with Ca removed define how dramatic conformational changes in the cytoplasmic domains may communicate with the VSD during activation. Together four structures provide a basis for understanding the voltage-dependent transition from resting to activated state, the tuning of VSD by thermodynamic stability, and this channel's requirement of cytoplasmic Ca ions for activation. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 6e1p.cif.gz | 255.5 KB | Display | PDBx/mmCIF format |
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PDB format | pdb6e1p.ent.gz | 213 KB | Display | PDB format |
PDBx/mmJSON format | 6e1p.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 6e1p_validation.pdf.gz | 1.3 MB | Display | wwPDB validaton report |
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Full document | 6e1p_full_validation.pdf.gz | 1.3 MB | Display | |
Data in XML | 6e1p_validation.xml.gz | 56.1 KB | Display | |
Data in CIF | 6e1p_validation.cif.gz | 80 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/e1/6e1p ftp://data.pdbj.org/pub/pdb/validation_reports/e1/6e1p | HTTPS FTP |
-Related structure data
Related structure data | 8960MC 8956C 8957C 8958C 6cx0C 6e1kC 6e1mC 6e1nC C: citing same article (ref.) M: map data used to model this data |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
#1: Protein | Mass: 84547.203 Da / Num. of mol.: 2 / Mutation: D230N, D444N, Q518E Source method: isolated from a genetically manipulated source Source: (gene. exp.) Arabidopsis thaliana (thale cress) / Gene: TPC1, CCH1, FOU2, At4g03560, F9H3.19, T5L23.5 / Production host: Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: Q94KI8 #2: Chemical | ChemComp-CA / #3: Chemical | ChemComp-PLM / #4: Water | ChemComp-HOH / | |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: AtTPC1(DDE) / Type: COMPLEX / Entity ID: #1 / Source: MULTIPLE SOURCES |
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Source (natural) | Organism: Arabidopsis thaliana (thale cress) |
Buffer solution | pH: 7.3 |
Specimen | Conc.: 0.8 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES / Details: Sample reconstituted into saposin A. |
Vitrification | Instrument: FEI VITROBOT MARK III / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 20 K |
-Electron microscopy imaging
Experimental equipment | Model: Tecnai Polara / Image courtesy: FEI Company |
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Microscopy | Model: FEI POLARA 300 |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 31000 X / Calibrated magnification: 41132 X / Nominal defocus max: 2000 nm / Nominal defocus min: 800 nm / Cs: 2 mm / C2 aperture diameter: 70 µm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN |
Image recording | Average exposure time: 0.2 sec. / Electron dose: 1 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 2 / Num. of real images: 3408 |
Image scans | Movie frames/image: 60 / Used frames/image: 2-60 |
-Processing
Software |
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EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 996035 | ||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C2 (2 fold cyclic) | ||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 67308 / Num. of class averages: 1 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||
Atomic model building | Protocol: RIGID BODY FIT / Space: REAL | ||||||||||||||||||||||||||||||||
Refinement | Stereochemistry target values: GeoStd + Monomer Library | ||||||||||||||||||||||||||||||||
Refine LS restraints |
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