+Open data
-Basic information
Entry | Database: PDB / ID: 7n4t | |||||||||
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Title | Low conductance mechanosensitive channel YnaI | |||||||||
Components | Low conductance mechanosensitive channel YnaI | |||||||||
Keywords | TRANSPORT PROTEIN / Low conductance mechanosensitive channel YnaI / NCMN / ion channel / membrane protein | |||||||||
Function / homology | Function and homology information mechanosensitive monoatomic ion channel activity / cellular response to osmotic stress / identical protein binding / plasma membrane Similarity search - Function | |||||||||
Biological species | Escherichia coli 'BL21-GoldpLysS AG' | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / negative staining / cryo EM / Resolution: 2.4 Å | |||||||||
Authors | Catalano, C. / Ben-Hail, D. / Qiu, W. / des Georges, A. / Guo, Y. | |||||||||
Funding support | United States, 2items
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Citation | Journal: Membranes (Basel) / Year: 2021 Title: Cryo-EM Structure of Mechanosensitive Channel YnaI Using SMA2000: Challenges and Opportunities. Authors: Claudio Catalano / Danya Ben-Hail / Weihua Qiu / Paul Blount / Amedee des Georges / Youzhong Guo / Abstract: Mechanosensitive channels respond to mechanical forces exerted on the cell membrane and play vital roles in regulating the chemical equilibrium within cells and their environment. High-resolution ...Mechanosensitive channels respond to mechanical forces exerted on the cell membrane and play vital roles in regulating the chemical equilibrium within cells and their environment. High-resolution structural information is required to understand the gating mechanisms of mechanosensitive channels. Protein-lipid interactions are essential for the structural and functional integrity of mechanosensitive channels, but detergents cannot maintain the crucial native lipid environment for purified mechanosensitive channels. Recently, detergent-free systems have emerged as alternatives for membrane protein structural biology. This report shows that while membrane-active polymer, SMA2000, could retain some native cell membrane lipids on the transmembrane domain of the mechanosensitive-like YnaI channel, the complete structure of the transmembrane domain of YnaI was not resolved. This reveals a significant limitation of SMA2000 or similar membrane-active copolymers. This limitation may come from the heterogeneity of the polymers and nonspecific interactions between the polymers and the relatively large hydrophobic pockets within the transmembrane domain of YnaI. However, this limitation offers development opportunities for detergent-free technology for challenging membrane proteins. | |||||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 7n4t.cif.gz | 292 KB | Display | PDBx/mmCIF format |
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PDB format | pdb7n4t.ent.gz | 236.3 KB | Display | PDB format |
PDBx/mmJSON format | 7n4t.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 7n4t_validation.pdf.gz | 1.2 MB | Display | wwPDB validaton report |
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Full document | 7n4t_full_validation.pdf.gz | 1.2 MB | Display | |
Data in XML | 7n4t_validation.xml.gz | 57 KB | Display | |
Data in CIF | 7n4t_validation.cif.gz | 79.4 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/n4/7n4t ftp://data.pdbj.org/pub/pdb/validation_reports/n4/7n4t | HTTPS FTP |
-Related structure data
Related structure data | 24177MC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
#1: Protein | Mass: 38792.344 Da / Num. of mol.: 7 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria) Strain: K12 / Gene: ynaI, b1330, JW1323 Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria) References: UniProt: P0AEB5 #2: Chemical | ChemComp-PEE / Has ligand of interest | Y | |
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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: Low conductance mechanosensitive channel YnaI / Type: CELL / Entity ID: #1 / Source: NATURAL |
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Source (natural) | Organism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria) |
Source (recombinant) | Organism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria) |
Buffer solution | pH: 7.8 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: YES / Vitrification applied: YES |
EM staining | Type: NEGATIVE / Material: 2 % uranil acetate |
Specimen support | Grid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3 |
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE-PROPANE / Humidity: 100 % / Chamber temperature: 277.15 K |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD |
Image recording | Average exposure time: 8 sec. / Electron dose: 56.18 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 2155 |
Image scans | Movie frames/image: 40 |
-Processing
Software | Name: PHENIX / Version: 1.19_4092: / Classification: refinement | ||||||||||||||||||||||||||||
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EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 330000 | ||||||||||||||||||||||||||||
Symmetry | Point symmetry: C7 (7 fold cyclic) | ||||||||||||||||||||||||||||
3D reconstruction | Resolution: 2.4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 142000 / Symmetry type: POINT | ||||||||||||||||||||||||||||
Atomic model building | Protocol: OTHER / Space: REAL | ||||||||||||||||||||||||||||
Atomic model building | PDB-ID: 5Y4O Accession code: 5Y4O / Source name: PDB / Type: experimental model | ||||||||||||||||||||||||||||
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