+Open data
-Basic information
Entry | Database: PDB / ID: 8gcl | ||||||
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Title | Cryo-EM structure of hAQP2 in DDM | ||||||
Components | Aquaporin-2 | ||||||
Keywords | MEMBRANE PROTEIN / channel | ||||||
Function / homology | Function and homology information cellular response to water deprivation / renal water transport / glycerol transmembrane transporter activity / water transmembrane transporter activity / Passive transport by Aquaporins / lumenal side of membrane / glycerol transmembrane transport / cellular response to mercury ion / water transport / water channel activity ...cellular response to water deprivation / renal water transport / glycerol transmembrane transporter activity / water transmembrane transporter activity / Passive transport by Aquaporins / lumenal side of membrane / glycerol transmembrane transport / cellular response to mercury ion / water transport / water channel activity / metanephric collecting duct development / renal water homeostasis / transport vesicle membrane / cellular response to copper ion / actin filament organization / recycling endosome / Vasopressin regulates renal water homeostasis via Aquaporins / basolateral plasma membrane / protein homotetramerization / apical plasma membrane / perinuclear region of cytoplasm / Golgi apparatus / extracellular exosome / membrane / plasma membrane Similarity search - Function | ||||||
Biological species | Homo sapiens (human) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.89 Å | ||||||
Authors | Kamegawa, A. / Suzuki, S. / Nishikawa, K. / Numoto, N. / Suzuki, H. / Fujiyoshi, Y. | ||||||
Funding support | Japan, 1items
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Citation | Journal: J Struct Biol / Year: 2023 Title: Structural analysis of the water channel AQP2 by single-particle cryo-EM. Authors: Akiko Kamegawa / Shota Suzuki / Hiroshi Suzuki / Kouki Nishikawa / Nobutaka Numoto / Yoshinori Fujiyoshi / Abstract: Water channels, which are small membrane proteins almost entirely buried in lipid membranes, are challenging research targets for single-particle cryo-electron microscopy (cryo-EM), a powerful ...Water channels, which are small membrane proteins almost entirely buried in lipid membranes, are challenging research targets for single-particle cryo-electron microscopy (cryo-EM), a powerful technique routinely used to determine the structures of membrane proteins. Because the single-particle method enables structural analysis of a whole protein with flexible parts that interfere with crystallization, we have focused our efforts on analyzing water channel structures. Here, utilizing this system, we analyzed the structure of full-length aquaporin-2 (AQP2), a primary regulator of vasopressin-dependent reabsorption of water at the renal collecting ducts. The 2.9 Å resolution map revealed a cytoplasmic extension of the cryo-EM density that was presumed to be the highly flexible C-terminus at which the localization of AQP2 is regulated in the renal collecting duct cells. We also observed a continuous density along the common water pathway inside the channel pore and lipid-like molecules at the membrane interface. Observations of these constructions in the AQP2 structure analyzed without any fiducial markers (e.g., a rigidly bound antibody) indicate that single-particle cryo-EM will be useful for investigating water channels in native states as well as in complexes with chemical compounds. | ||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8gcl.cif.gz | 56.9 KB | Display | PDBx/mmCIF format |
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PDB format | pdb8gcl.ent.gz | 38.7 KB | Display | PDB format |
PDBx/mmJSON format | 8gcl.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 8gcl_validation.pdf.gz | 1.3 MB | Display | wwPDB validaton report |
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Full document | 8gcl_full_validation.pdf.gz | 1.3 MB | Display | |
Data in XML | 8gcl_validation.xml.gz | 19.6 KB | Display | |
Data in CIF | 8gcl_validation.cif.gz | 27.5 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/gc/8gcl ftp://data.pdbj.org/pub/pdb/validation_reports/gc/8gcl | HTTPS FTP |
-Related structure data
Related structure data | 29934MC 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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Symmetry | Point symmetry: (Schoenflies symbol: C4 (4 fold cyclic)) | ||||||||||||||||||||
Noncrystallographic symmetry (NCS) | NCS oper:
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-Components
#1: Protein | Mass: 28862.389 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: AQP2 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P41181 |
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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: tetrameter of hAQP2 / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT |
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Molecular weight | Experimental value: NO |
Source (natural) | Organism: Homo sapiens (human) |
Source (recombinant) | Organism: Spodoptera frugiperda (fall armyworm) |
Buffer solution | pH: 8 |
Specimen | Conc.: 7.2 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Specimen support | Grid material: MOLYBDENUM / Grid mesh size: 200 divisions/in. / Grid type: Quantifoil R1.2/1.3 |
Vitrification | Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 295 K |
-Electron microscopy imaging
Microscopy | Model: JEOL CRYO ARM 300 |
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Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 2500 nm / Nominal defocus min: 1200 nm |
Image recording | Electron dose: 69.6 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) |
-Processing
Software | Name: REFMAC / Version: 5.8.0352 / Classification: refinement | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 2.89 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 236700 / Symmetry type: POINT | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Refinement | Resolution: 2.89→2.89 Å / Cor.coef. Fo:Fc: 0.838 / SU B: 11.101 / SU ML: 0.207 / ESU R: 0.194 Stereochemistry target values: MAXIMUM LIKELIHOOD WITH PHASES Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS
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Solvent computation | Solvent model: PARAMETERS FOR MASK CACLULATION | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Displacement parameters | Biso mean: 91.506 Å2 | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Refinement step | Cycle: 1 / Total: 1754 | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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