[English] 日本語
Yorodumi- PDB-7ph3: AMP-PNP bound nanodisc reconstituted MsbA with nanobodies, spin-l... -
+Open data
-Basic information
Entry | Database: PDB / ID: 7ph3 | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Title | AMP-PNP bound nanodisc reconstituted MsbA with nanobodies, spin-labeled at position A60C | ||||||||||||
Components |
| ||||||||||||
Keywords | MEMBRANE PROTEIN / ABC transporter / nanobody / AMP-PNP / Gd-DOTA | ||||||||||||
Function / homology | Function and homology information MsbA transporter complex / lipopolysaccharide floppase activity / lipid translocation / ABC-type lipid A-core oligosaccharide transporter / lipopolysaccharide transport / ATPase-coupled lipid transmembrane transporter activity / ABC-type xenobiotic transporter activity / lipid transport / ATP-binding cassette (ABC) transporter complex / transmembrane transport ...MsbA transporter complex / lipopolysaccharide floppase activity / lipid translocation / ABC-type lipid A-core oligosaccharide transporter / lipopolysaccharide transport / ATPase-coupled lipid transmembrane transporter activity / ABC-type xenobiotic transporter activity / lipid transport / ATP-binding cassette (ABC) transporter complex / transmembrane transport / lipid binding / ATP hydrolysis activity / ATP binding / membrane / identical protein binding / plasma membrane Similarity search - Function | ||||||||||||
Biological species | Escherichia coli K-12 (bacteria) Vicugna pacos (alpaca) | ||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.8 Å | ||||||||||||
Authors | Parey, K. / Januliene, D. / Galazzo, L. / Meier, G. / Vecchis, D. / Striednig, B. / Hilbi, H. / Schaefer, L.V. / Kuprov, I. / Bordignon, E. ...Parey, K. / Januliene, D. / Galazzo, L. / Meier, G. / Vecchis, D. / Striednig, B. / Hilbi, H. / Schaefer, L.V. / Kuprov, I. / Bordignon, E. / Seeger, M.A. / Moeller, A. | ||||||||||||
Funding support | Germany, 3items
| ||||||||||||
Citation | Journal: Sci Adv / Year: 2022 Title: The ABC transporter MsbA adopts the wide inward-open conformation in cells. Authors: Laura Galazzo / Gianmarco Meier / Dovile Januliene / Kristian Parey / Dario De Vecchis / Bianca Striednig / Hubert Hilbi / Lars V Schäfer / Ilya Kuprov / Arne Moeller / Enrica Bordignon / Markus A Seeger / Abstract: Membrane proteins are currently investigated after detergent extraction from native cellular membranes and reconstitution into artificial liposomes or nanodiscs, thereby removing them from their ...Membrane proteins are currently investigated after detergent extraction from native cellular membranes and reconstitution into artificial liposomes or nanodiscs, thereby removing them from their physiological environment. However, to truly understand the biophysical properties of membrane proteins in a physiological environment, they must be investigated within living cells. Here, we used a spin-labeled nanobody to interrogate the conformational cycle of the ABC transporter MsbA by double electron-electron resonance. Unexpectedly, the wide inward-open conformation of MsbA, commonly considered a nonphysiological state, was found to be prominently populated in cells. Molecular dynamics simulations revealed that extensive lateral portal opening is essential to provide access of its large natural substrate core lipid A to the binding cavity. Our work paves the way to investigate the conformational landscape of membrane proteins in cells. | ||||||||||||
History |
|
-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
---|
-Downloads & links
-Download
PDBx/mmCIF format | 7ph3.cif.gz | 248 KB | Display | PDBx/mmCIF format |
---|---|---|---|---|
PDB format | pdb7ph3.ent.gz | 203.2 KB | Display | PDB format |
PDBx/mmJSON format | 7ph3.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/ph/7ph3 ftp://data.pdbj.org/pub/pdb/validation_reports/ph/7ph3 | HTTPS FTP |
---|
-Related structure data
Related structure data | 13405MC 7ndfC 7ph2C 7ph4C 7ph7C M: map data used to model this data C: citing same article (ref.) |
---|---|
Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
|
---|---|
1 |
|
-Components
-Protein / Antibody / Sugars , 3 types, 6 molecules ABCD
#1: Protein | Mass: 65760.852 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Strain: K12 / Gene: msbA, b0914, JW0897 / Production host: Escherichia coli (E. coli) References: UniProt: P60752, ABC-type lipid A-core oligosaccharide transporter #2: Antibody | Mass: 12440.957 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Vicugna pacos (alpaca) / Production host: Escherichia coli (E. coli) #5: Sugar | |
---|
-Non-polymers , 6 types, 14 molecules
#3: Chemical | #4: Chemical | #6: Chemical | #7: Chemical | #8: Chemical | #9: Water | ChemComp-HOH / | |
---|
-Details
Has ligand of interest | Y |
---|
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
---|---|
EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: E. coli MsbA in complex with nanobody Nb_MsbA#1, labeled with Gd-DOTA at Cys60 Type: COMPLEX / Entity ID: #1-#2 / Source: RECOMBINANT |
---|---|
Source (natural) | Organism: Escherichia coli (E. coli) |
Source (recombinant) | Organism: Escherichia coli (E. coli) |
Buffer solution | pH: 7.5 |
Specimen | Conc.: 2.6 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Cryogen name: ETHANE |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
---|---|
Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2500 nm / Nominal defocus min: 800 nm |
Image recording | Electron dose: 50 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) |
-Processing
EM software |
| ||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||
Symmetry | Point symmetry: C2 (2 fold cyclic) | ||||||||||||||||
3D reconstruction | Resolution: 2.8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 109465 / Symmetry type: POINT | ||||||||||||||||
Refinement | Highest resolution: 2.8 Å |