+
Open data
-
Basic information
Entry | Database: PDB / ID: 8i8r | ||||||
---|---|---|---|---|---|---|---|
Title | Cryo-EM Structure of OmpC3-MlaA Complex in MSP2N2 Nanodiscs | ||||||
![]() |
| ||||||
![]() | LIPID TRANSPORT / bacteria / outer membrane / phospholipid / lipid asymmetry / membrane protein / protein complex structure / channel | ||||||
Function / homology | ![]() intermembrane phospholipid transfer / porin activity / pore complex / monoatomic ion transmembrane transport / cell outer membrane / virus receptor activity / receptor-mediated virion attachment to host cell / DNA damage response / identical protein binding / metal ion binding Similarity search - Function | ||||||
Biological species | ![]() ![]() | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.93 Å | ||||||
![]() | Yeow, J. / Luo, M. / Chng, S.S. | ||||||
Funding support | ![]()
| ||||||
![]() | ![]() Title: Molecular mechanism of phospholipid transport at the bacterial outer membrane interface. Authors: Jiang Yeow / Min Luo / Shu-Sin Chng / ![]() Abstract: The outer membrane (OM) of Gram-negative bacteria is an asymmetric lipid bilayer with outer leaflet lipopolysaccharides and inner leaflet phospholipids (PLs). This unique lipid asymmetry renders the ...The outer membrane (OM) of Gram-negative bacteria is an asymmetric lipid bilayer with outer leaflet lipopolysaccharides and inner leaflet phospholipids (PLs). This unique lipid asymmetry renders the OM impermeable to external insults, including antibiotics and bile salts. To maintain this barrier, the OmpC-Mla system removes mislocalized PLs from the OM outer leaflet, and transports them to the inner membrane (IM); in the first step, the OmpC-MlaA complex transfers PLs to the periplasmic chaperone MlaC, but mechanistic details are lacking. Here, we biochemically and structurally characterize the MlaA-MlaC transient complex. We map the interaction surfaces between MlaA and MlaC in Escherichia coli, and show that electrostatic interactions are important for MlaC recruitment to the OM. We further demonstrate that interactions with MlaC modulate conformational states in MlaA. Finally, we solve a 2.9-Å cryo-EM structure of a disulfide-trapped OmpC-MlaA-MlaC complex in nanodiscs, reinforcing the mechanism of MlaC recruitment, and highlighting membrane thinning as a plausible strategy for directing lipids for transport. Our work offers critical insights into retrograde PL transport by the OmpC-Mla system in maintaining OM lipid asymmetry. | ||||||
History |
|
-
Structure visualization
Structure viewer | Molecule: ![]() ![]() |
---|
-
Downloads & links
-
Download
PDBx/mmCIF format | ![]() | 261.1 KB | Display | ![]() |
---|---|---|---|---|
PDB format | ![]() | Display | ![]() | |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Arichive directory | ![]() ![]() | HTTPS FTP |
---|
-Related structure data
Related structure data | ![]() 35250MC ![]() 8i8xC M: map data used to model this data C: citing same article ( |
---|---|
Similar structure data | Similarity search - Function & homology ![]() |
-
Links
-
Assembly
Deposited unit | ![]()
|
---|---|
1 |
|
-
Components
#1: Protein | Mass: 38336.242 Da / Num. of mol.: 3 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() #2: Protein | | Mass: 26298.336 Da / Num. of mol.: 1 / Mutation: Q205C Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() #3: Chemical | Has ligand of interest | N | |
---|
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
---|---|
EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-
Sample preparation
Component |
| ||||||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Molecular weight |
| ||||||||||||||||||||||||||||
Source (natural) |
| ||||||||||||||||||||||||||||
Source (recombinant) |
| ||||||||||||||||||||||||||||
Buffer solution | pH: 8 Details: Tris-buffered saline (TBS) buffer (20 mM Tris HCl pH 8.0, 150 mM NaCl) | ||||||||||||||||||||||||||||
Buffer component |
| ||||||||||||||||||||||||||||
Specimen | Conc.: 12 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||||||||||
Specimen support | Grid material: COPPER / Grid mesh size: 200 divisions/in. / Grid type: Quantifoil R1.2/1.3 | ||||||||||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K |
-
Electron microscopy imaging
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
---|---|
Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: ![]() |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 105000 X / Nominal defocus max: 2000 nm / Nominal defocus min: 800 nm / Cs: 2.7 mm |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 5.99 sec. / Electron dose: 90 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 6018 Details: Images were collected in movie-mode at 50 frames per image |
EM imaging optics | Energyfilter name: GIF Tridiem 4K Details: Gatan GIF post-column energy filter operated in zero-loss mode Energyfilter slit width: 20 eV |
Image scans | Sampling size: 5.2 µm / Width: 5760 / Height: 4092 |
-
Processing
EM software |
| ||||||||||||||||||||||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
CTF correction | Details: Patch CTF routine in cryoSPARC / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 2434319 Details: We performed automated particle picking using Blob and Template Picker for initial template picking and final sampling respectively. | ||||||||||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 2.93 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 205487 / Algorithm: BACK PROJECTION / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | B value: 88 / Protocol: FLEXIBLE FIT / Space: REAL / Target criteria: Cross-correlation coefficient Details: Building of the high-resolution structures of OmpC-MlaA was done via approximate fitting using the OmpC-MlaA crystal structure (PDB 5NUP) as rigid bodies into the density in Chimera, ...Details: Building of the high-resolution structures of OmpC-MlaA was done via approximate fitting using the OmpC-MlaA crystal structure (PDB 5NUP) as rigid bodies into the density in Chimera, followed by refinement in COOT. | ||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | 3D fitting-ID: 1 / Accession code: 5NUP / Details: The initial model consisted of the chains A,B,C,D for PDB entry 5NUP / Initial refinement model-ID: 1 / PDB-ID: 5NUP / Source name: PDB / Type: experimental model
| ||||||||||||||||||||||||||||||||||||||||||||
Refine LS restraints |
|