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- PDB-7ch0: The overall structure of the MlaFEDB complex in ATP-bound EQclose... -

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Basic information

Entry
Database: PDB / ID: 7ch0
TitleThe overall structure of the MlaFEDB complex in ATP-bound EQclose conformation (Mutation of E170Q on MlaF)
Components
  • Lipid asymmetry maintenance ABC transporter permease subunit MlaE
  • Lipid asymmetry maintenance protein MlaB
  • Outer membrane lipid asymmetry maintenance protein MlaD
  • Phospholipid ABC transporter ATP-binding protein MlaF
KeywordsMEMBRANE PROTEIN
Function / homology
Function and homology information


phospholipid transfer activity / intermembrane phospholipid transfer / phospholipid transporter activity / phospholipid-translocating ATPase complex / phospholipid transport / Translocases; Catalysing the translocation of other compounds; Linked to the hydrolysis of a nucleoside triphosphate / ATPase-coupled transmembrane transporter activity / ATP-binding cassette (ABC) transporter complex / phospholipid binding / response to antibiotic ...phospholipid transfer activity / intermembrane phospholipid transfer / phospholipid transporter activity / phospholipid-translocating ATPase complex / phospholipid transport / Translocases; Catalysing the translocation of other compounds; Linked to the hydrolysis of a nucleoside triphosphate / ATPase-coupled transmembrane transporter activity / ATP-binding cassette (ABC) transporter complex / phospholipid binding / response to antibiotic / DNA damage response / ATP hydrolysis activity / ATP binding / membrane / plasma membrane / cytosol
Similarity search - Function
Probable ABC transporter ATP-binding protein MlaF/Mkl / : / : / : / Probable phospholipid ABC transporter-binding protein MlaD / ABC transport permease subunit MlaE, Proteobacteria / : / ABC transporter permease MalE / Permease MlaE / STAS domain ...Probable ABC transporter ATP-binding protein MlaF/Mkl / : / : / : / Probable phospholipid ABC transporter-binding protein MlaD / ABC transport permease subunit MlaE, Proteobacteria / : / ABC transporter permease MalE / Permease MlaE / STAS domain / STAS domain / Mce/MlaD / MlaD protein / Transcription Regulator spoIIAA / STAS domain profile. / STAS domain / STAS domain superfamily / ABC transporter-like, conserved site / ABC transporters family signature. / ABC transporter / ABC transporter-like, ATP-binding domain / ATP-binding cassette, ABC transporter-type domain profile. / P-loop containing nucleotide triphosphate hydrolases / ATPases associated with a variety of cellular activities / AAA+ ATPase domain / Rossmann fold / P-loop containing nucleoside triphosphate hydrolase / 2-Layer Sandwich / 3-Layer(aba) Sandwich / Alpha Beta
Similarity search - Domain/homology
ADENOSINE-5'-TRIPHOSPHATE / Intermembrane phospholipid transport system permease protein MlaE / Lipid asymmetry maintenance protein MlaB / Phospholipid ABC transporter ATP-binding protein MlaF / Outer membrane lipid asymmetry maintenance protein MlaD / Intermembrane phospholipid transport system ATP-binding protein MlaF / Intermembrane phospholipid transport system binding protein MlaB / Intermembrane phospholipid transport system binding protein MlaD / Intermembrane phospholipid transport system permease protein MlaE
Similarity search - Component
Biological speciesEscherichia coli K-12 (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.7 Å
AuthorsChi, X.M. / Fan, Q.X. / Zhang, Y.Y. / Liang, K. / Zhou, Q. / Li, Y.Y.
Funding support China, 1items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)31930059 China
CitationJournal: Cell Res / Year: 2020
Title: Structural mechanism of phospholipids translocation by MlaFEDB complex.
Authors: Ximin Chi / Qiongxuan Fan / Yuanyuan Zhang / Ke Liang / Li Wan / Qiang Zhou / Yanyan Li /
Abstract: In Gram-negative bacteria, phospholipids are major components of the inner membrane and the inner leaflet of the outer membrane, playing an essential role in forming the unique dual-membrane barrier ...In Gram-negative bacteria, phospholipids are major components of the inner membrane and the inner leaflet of the outer membrane, playing an essential role in forming the unique dual-membrane barrier to exclude the entry of most antibiotics. Understanding the mechanisms of phospholipid translocation between the inner and outer membrane represents one of the major challenges surrounding bacterial phospholipid homeostasis. The conserved MlaFEDB complex in the inner membrane functions as an ABC transporter to drive the translocation of phospholipids between the inner membrane and the periplasmic protein MlaC. However, the mechanism of phospholipid translocation remains elusive. Here we determined three cryo-EM structures of MlaFEDB from Escherichia coli in its nucleotide-free and ATP-bound conformations, and performed extensive functional studies to verify and extend our findings from structural analyses. Our work reveals unique structural features of the entire MlaFEDB complex, six well-resolved phospholipids in three distinct cavities, and large-scale conformational changes upon ATP binding. Together, these findings define the cycle of structural rearrangement of MlaFEDB in action, and suggest that MlaFEDB uses an extrusion mechanism to extract and release phospholipids through the central translocation cavity.
History
DepositionJul 3, 2020Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Sep 9, 2020Provider: repository / Type: Initial release
Revision 1.1Sep 16, 2020Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year
Revision 2.0Nov 18, 2020Group: Atomic model / Data collection ...Atomic model / Data collection / Derived calculations / Source and taxonomy
Category: atom_site / em_software ...atom_site / em_software / entity_src_gen / pdbx_struct_assembly_prop / pdbx_struct_sheet_hbond / pdbx_validate_torsion / struct_conf / struct_sheet / struct_sheet_order / struct_sheet_range / struct_site / struct_site_gen
Item: _atom_site.B_iso_or_equiv / _atom_site.Cartn_x ..._atom_site.B_iso_or_equiv / _atom_site.Cartn_x / _atom_site.Cartn_y / _atom_site.Cartn_z / _em_software.category / _entity_src_gen.gene_src_strain / _entity_src_gen.pdbx_gene_src_scientific_name / _entity_src_gen.pdbx_host_org_scientific_name / _entity_src_gen.pdbx_host_org_strain / _struct_site.pdbx_num_residues
Description: Model orientation/position / Provider: author / Type: Coordinate replacement
Revision 2.1Dec 23, 2020Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation_author.identifier_ORCID
Revision 2.2Mar 27, 2024Group: Data collection / Database references / Category: chem_comp_atom / chem_comp_bond / database_2
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession

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Structure visualization

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Assembly

Deposited unit
A: Lipid asymmetry maintenance ABC transporter permease subunit MlaE
B: Phospholipid ABC transporter ATP-binding protein MlaF
E: Phospholipid ABC transporter ATP-binding protein MlaF
D: Lipid asymmetry maintenance ABC transporter permease subunit MlaE
C: Lipid asymmetry maintenance protein MlaB
F: Lipid asymmetry maintenance protein MlaB
G: Outer membrane lipid asymmetry maintenance protein MlaD
H: Outer membrane lipid asymmetry maintenance protein MlaD
I: Outer membrane lipid asymmetry maintenance protein MlaD
J: Outer membrane lipid asymmetry maintenance protein MlaD
K: Outer membrane lipid asymmetry maintenance protein MlaD
L: Outer membrane lipid asymmetry maintenance protein MlaD
hetero molecules


Theoretical massNumber of molelcules
Total (without water)253,98014
Polymers252,96512
Non-polymers1,0142
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: microscopy, cryo EM map of the MlaFEDB complex in ATP-bound EQclose conformation (Mutation of E170Q on MlaF)
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein Lipid asymmetry maintenance ABC transporter permease subunit MlaE


Mass: 27885.162 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Strain: K-12 / Gene: mlaE, FAZ83_04790 / Production host: Escherichia coli K-12 (bacteria) / Strain (production host): K-12 / References: UniProt: A0A4S5B3V0, UniProt: P64606*PLUS
#2: Protein Phospholipid ABC transporter ATP-binding protein MlaF


Mass: 29127.816 Da / Num. of mol.: 2 / Mutation: E170Q
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Strain: K-12 / Gene: mlaF, FAZ83_04795 / Production host: Escherichia coli K-12 (bacteria) / Strain (production host): K-12 / References: UniProt: A0A4V3YUQ9, UniProt: P63386*PLUS
#3: Protein Lipid asymmetry maintenance protein MlaB


Mass: 10690.313 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Strain: K-12 / Gene: mlaB, FAZ83_04775 / Production host: Escherichia coli K-12 (bacteria) / Strain (production host): K-12 / References: UniProt: A0A4S5B5E3, UniProt: P64602*PLUS
#4: Protein
Outer membrane lipid asymmetry maintenance protein MlaD


Mass: 19593.133 Da / Num. of mol.: 6
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Strain: K-12 / Gene: mlaD, FAZ83_04785 / Production host: Escherichia coli K-12 (bacteria) / Strain (production host): K-12 / References: UniProt: A0A6D2XU65, UniProt: P64604*PLUS
#5: Chemical ChemComp-ATP / ADENOSINE-5'-TRIPHOSPHATE


Mass: 507.181 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C10H16N5O13P3 / Feature type: SUBJECT OF INVESTIGATION / Comment: ATP, energy-carrying molecule*YM
Has ligand of interestY

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: cryo EM map of the MlaFEDB complex in ATP-bound EQclose conformation (Mutation of E170Q on MlaF)
Type: COMPLEX / Entity ID: #1-#4 / Source: RECOMBINANT
Source (natural)Organism: Escherichia coli K-12 (bacteria)
Source (recombinant)Organism: Escherichia coli K-12 (bacteria)
Buffer solutionpH: 8
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

EM softwareName: RELION / Version: 3.0.6 / Category: 3D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 21159 / Symmetry type: POINT

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