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Open data
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Basic information
Entry | Database: PDB / ID: 7ch7 | ||||||
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Title | Cryo-EM structure of E.coli MlaFEB | ||||||
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![]() | MEMBRANE PROTEIN / Mla complex / Lipid transporter | ||||||
Function / homology | ![]() ATPase-coupled transmembrane transporter activity / ATP-binding cassette (ABC) transporter complex / ATP binding Similarity search - Function | ||||||
Biological species | ![]() ![]() | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.9 Å | ||||||
![]() | Zhou, C. / Shi, H. / Huang, Y. | ||||||
![]() | ![]() Title: Structural Insight into Phospholipid Transport by the MlaFEBD Complex from P. aeruginosa. Authors: Changping Zhou / Huigang Shi / Manfeng Zhang / Lijun Zhou / Le Xiao / Shasha Feng / Wonpil Im / Min Zhou / Xinzheng Zhang / Yihua Huang / ![]() ![]() Abstract: The outer membrane (OM) of Gram-negative bacteria, which consists of lipopolysaccharides (LPS) in the outer leaflet and phospholipids (PLs) in the inner leaflet, plays a key role in antibiotic ...The outer membrane (OM) of Gram-negative bacteria, which consists of lipopolysaccharides (LPS) in the outer leaflet and phospholipids (PLs) in the inner leaflet, plays a key role in antibiotic resistance and pathogen virulence. The maintenance of lipid asymmetry (Mla) pathway is known to be involved in PL transport and contributes to the lipid homeostasis of the OM, yet the underlying molecular mechanism and the directionality of PL transport in this pathway remain elusive. Here, we reported the cryo-EM structures of the ATP-binding cassette (ABC) transporter MlaFEBD from P. areuginosa, the core complex in the Mla pathway, in nucleotide-free (apo)-, ADP (ATP + vanadate)- and ATP (AMPPNP)-bound states as well as the structures of MlaFEB from E. coli in apo- and AMPPNP-bound states at a resolution range of 3.4-3.9 Å. The structures show that the MlaFEBD complex contains a total of twelve protein molecules with a stoichiometry of MlaFEBD, and binds a plethora of PLs at different locations. In contrast to canonical ABC transporters, nucleotide binding fails to trigger significant conformational changes of both MlaFEBD and MlaFEB in the nucleotide-binding and transmembrane domains of the ABC transporter, correlated with their low ATPase activities exhibited in both detergent micelles and lipid nanodiscs. Intriguingly, PLs or detergents appeared to relocate to the membrane-proximal end from the distal end of the hydrophobic tunnel formed by the MlaD hexamer in MlaFEBD upon addition of ATP, indicating that retrograde PL transport might occur in the tunnel in an ATP-dependent manner. Site-specific photocrosslinking experiment confirms that the substrate-binding pocket in the dimeric MlaE and the MlaD hexamer are able to bind PLs in vitro, in line with the notion that MlaFEBD complex functions as a PL transporter. | ||||||
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Structure visualization
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Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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PDBx/mmCIF format | ![]() | 206.7 KB | Display | ![]() |
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PDB format | ![]() | 167.2 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
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-Validation report
Arichive directory | ![]() ![]() | HTTPS FTP |
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-Related structure data
Related structure data | ![]() 30370MC ![]() 7ch6C ![]() 7ch8C ![]() 7ch9C ![]() 7chaC M: map data used to model this data C: citing same article ( |
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Similar structure data |
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Links
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Assembly
Deposited unit | ![]()
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Components
#1: Protein | Mass: 27885.162 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Strain: K12 / Gene: mlaE, FAZ83_04790 / Production host: ![]() ![]() #2: Protein | Mass: 29128.801 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Strain: K12 / Gene: mlaF, FAZ83_04795 / Production host: ![]() ![]() #3: Protein | Mass: 10690.313 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Strain: K12 / Gene: mlaB, FAZ83_04775 / Production host: ![]() ![]() |
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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 |
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Sample preparation
Component | Name: E.coli Mla FEB complex / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT |
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Molecular weight | Experimental value: YES |
Source (natural) | Organism: ![]() ![]() |
Source (recombinant) | Organism: ![]() ![]() |
Buffer solution | pH: 8 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Cryogen name: ETHANE |
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Electron microscopy imaging
Experimental equipment | ![]() Model: Talos Arctica / Image courtesy: FEI Company |
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Microscopy | Model: FEI TALOS ARCTICA |
Electron gun | Electron source: ![]() |
Electron lens | Mode: DIFFRACTION / Cs: 2.7 mm / C2 aperture diameter: 50 µm |
Specimen holder | Cryogen: NITROGEN |
Image recording | Electron dose: 50 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K2 QUANTUM (4k x 4k) |
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Processing
Software | Name: PHENIX / Version: 1.17.1_3660: / Classification: refinement | ||||||||||||||||||||||||||||
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EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||
Symmetry | Point symmetry: C2 (2 fold cyclic) | ||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.9 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 74479 / Symmetry type: POINT | ||||||||||||||||||||||||||||
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