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- PDB-8t3k: Heterodimeric ABC transporter BmrCD in the inward-facing conforma... -

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Basic information

Entry
Database: PDB / ID: 8t3k
TitleHeterodimeric ABC transporter BmrCD in the inward-facing conformation bound to ATP: BmrCD_IF-ATP2
Components
  • Probable multidrug resistance ABC transporter ATP-binding/permease protein YheH
  • Probable multidrug resistance ABC transporter ATP-binding/permease protein YheI
KeywordsTRANSLOCASE / transporter / complex / lipids / MEMBRANE PROTEIN
Function / homology
Function and homology information


ATPase-coupled lipid transmembrane transporter activity / Translocases; Catalysing the translocation of other compounds; Linked to the hydrolysis of a nucleoside triphosphate / ATPase-coupled transmembrane transporter activity / ABC-type transporter activity / transmembrane transport / response to antibiotic / ATP hydrolysis activity / ATP binding / plasma membrane
Similarity search - Function
Type 1 protein exporter / ABC transporter transmembrane region / ABC transporter type 1, transmembrane domain / ABC transporter integral membrane type-1 fused domain profile. / ABC transporter type 1, transmembrane 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. ...Type 1 protein exporter / ABC transporter transmembrane region / ABC transporter type 1, transmembrane domain / ABC transporter integral membrane type-1 fused domain profile. / ABC transporter type 1, transmembrane 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. / ATPases associated with a variety of cellular activities / AAA+ ATPase domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
ADENOSINE-5'-TRIPHOSPHATE / Chem-POV / Probable multidrug resistance ABC transporter ATP-binding/permease protein YheH / Probable multidrug resistance ABC transporter ATP-binding/permease protein YheI
Similarity search - Component
Biological speciesBacillus subtilis subsp. subtilis str. 168 (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.33 Å
AuthorsTang, Q. / Mchaourab, H.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)NIGMS R01-128087 United States
CitationJournal: Nat Commun / Year: 2023
Title: Asymmetric conformations and lipid interactions shape the ATP-coupled cycle of a heterodimeric ABC transporter.
Authors: Qingyu Tang / Matt Sinclair / Hale S Hasdemir / Richard A Stein / Erkan Karakas / Emad Tajkhorshid / Hassane S Mchaourab /
Abstract: Here we used cryo-electron microscopy (cryo-EM), double electron-electron resonance spectroscopy (DEER), and molecular dynamics (MD) simulations, to capture and characterize ATP- and substrate-bound ...Here we used cryo-electron microscopy (cryo-EM), double electron-electron resonance spectroscopy (DEER), and molecular dynamics (MD) simulations, to capture and characterize ATP- and substrate-bound inward-facing (IF) and occluded (OC) conformational states of the heterodimeric ATP binding cassette (ABC) multidrug exporter BmrCD in lipid nanodiscs. Supported by DEER analysis, the structures reveal that ATP-powered isomerization entails changes in the relative symmetry of the BmrC and BmrD subunits that propagates from the transmembrane domain to the nucleotide binding domain. The structures uncover asymmetric substrate and Mg binding which we hypothesize are required for triggering ATP hydrolysis preferentially in one of the nucleotide-binding sites. MD simulations demonstrate that multiple lipid molecules differentially bind the IF versus the OC conformation thus establishing that lipid interactions modulate BmrCD energy landscape. Our findings are framed in a model that highlights the role of asymmetric conformations in the ATP-coupled transport with general implications to the mechanism of ABC transporters.
History
DepositionJun 7, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0Nov 29, 2023Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
D: Probable multidrug resistance ABC transporter ATP-binding/permease protein YheH
C: Probable multidrug resistance ABC transporter ATP-binding/permease protein YheI
hetero molecules


Theoretical massNumber of molelcules
Total (without water)155,86817
Polymers144,9732
Non-polymers10,89515
Water0
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein Probable multidrug resistance ABC transporter ATP-binding/permease protein YheH


Mass: 77369.898 Da / Num. of mol.: 1 / Mutation: C154A,C256A,C351A,E592Q
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Bacillus subtilis subsp. subtilis str. 168 (bacteria)
Gene: yheH, BSU09720 / Production host: Escherichia coli (E. coli)
References: UniProt: O07549, Translocases; Catalysing the translocation of other compounds; Linked to the hydrolysis of a nucleoside triphosphate
#2: Protein Probable multidrug resistance ABC transporter ATP-binding/permease protein YheI


Mass: 67602.961 Da / Num. of mol.: 1 / Mutation: D500Q
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Bacillus subtilis subsp. subtilis str. 168 (bacteria)
Gene: yheI, BSU09710 / Production host: Escherichia coli (E. coli)
References: UniProt: O07550, Translocases; Catalysing the translocation of other compounds; Linked to the hydrolysis of a nucleoside triphosphate
#3: Chemical
ChemComp-POV / (2S)-3-(hexadecanoyloxy)-2-[(9Z)-octadec-9-enoyloxy]propyl 2-(trimethylammonio)ethyl phosphate / POPC / POPC


Mass: 760.076 Da / Num. of mol.: 13 / Source method: obtained synthetically / Formula: C42H82NO8P / Feature type: SUBJECT OF INVESTIGATION / Comment: phospholipid*YM
#4: Chemical ChemComp-ATP / ADENOSINE-5'-TRIPHOSPHATE / Adenosine 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: Heterodimeric BmrCD with ligands ATP and lipids / Type: COMPLEX / Entity ID: #1-#2 / Source: RECOMBINANT
Source (natural)Organism: Bacillus subtilis subsp. subtilis str. 168 (bacteria)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 8
SpecimenConc.: 3 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K

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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 FIELDBright-field microscopy / Nominal defocus max: 2200 nm / Nominal defocus min: 400 nm / Cs: 2.7 mm
Specimen holderCryogen: NITROGEN
Image recordingElectron dose: 15 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

CTF correctionType: NONE
3D reconstructionResolution: 3.33 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 76190 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00610215
ELECTRON MICROSCOPYf_angle_d0.913715
ELECTRON MICROSCOPYf_dihedral_angle_d18.3773936
ELECTRON MICROSCOPYf_chiral_restr0.0461511
ELECTRON MICROSCOPYf_plane_restr0.0061687

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