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- EMDB-41058: Heterodimeric ABC transporter BmrCD in the inward-facing conforma... -

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Basic information

Entry
Database: EMDB / ID: EMD-41058
TitleHeterodimeric ABC transporter BmrCD in the inward-facing conformation bound to substrate and ADPVi: BmrCD_IF-HT/ADPVi
Map data
Sample
  • Complex: Heterodimeric BmrCD with ligands ATP/AOV and lipids
    • Protein or peptide: Probable multidrug resistance ABC transporter ATP-binding/permease protein YheI
    • Protein or peptide: Probable multidrug resistance ABC transporter ATP-binding/permease protein YheH
Keywordstransporter / lipids / MEMBRANE PROTEIN / TRANSLOCASE
Biological speciesBacillus subtilis subsp. subtilis str. 168 (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 5.65 Å
AuthorsQingyu T / Hassane M
Funding support United States, 1 items
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 14, 2023-
Header (metadata) releaseNov 15, 2023-
Map releaseNov 15, 2023-
UpdateNov 29, 2023-
Current statusNov 29, 2023Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_41058.png

Downloads & links

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Map

FileDownload / File: emd_41058.map.gz / Format: CCP4 / Size: 421.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.65 Å/pix.
x 480 pix.
= 310.56 Å		0.65 Å/pix.
x 480 pix.
= 310.56 Å		0.65 Å/pix.
x 480 pix.
= 310.56 Å

Surface

Projections

Slices (1/3)

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Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.647 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.05
Minimum - Maximum-0.15549237 - 0.22112297
Average (Standard dev.)-0.00021095172 (±0.006516455)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions480480480
Spacing480480480
CellA=B=C: 310.56 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_41058_half_map_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_41058_half_map_2.map
Projections & Slices
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Histogram

Histogram (log scale)

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Sample components

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Entire : Heterodimeric BmrCD with ligands ATP/AOV and lipids

EntireName: Heterodimeric BmrCD with ligands ATP/AOV and lipids
Components
  • Complex: Heterodimeric BmrCD with ligands ATP/AOV and lipids
    • Protein or peptide: Probable multidrug resistance ABC transporter ATP-binding/permease protein YheI
    • Protein or peptide: Probable multidrug resistance ABC transporter ATP-binding/permease protein YheH

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Supramolecule #1: Heterodimeric BmrCD with ligands ATP/AOV and lipids

SupramoleculeName: Heterodimeric BmrCD with ligands ATP/AOV and lipids / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Bacillus subtilis subsp. subtilis str. 168 (bacteria)

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Macromolecule #1: Probable multidrug resistance ABC transporter ATP-binding/permeas...

MacromoleculeName: Probable multidrug resistance ABC transporter ATP-binding/permease protein YheI
type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
EC number: Translocases; Catalysing the translocation of other compounds; Linked to the hydrolysis of a nucleoside triphosphate
Source (natural)Organism: Bacillus subtilis subsp. subtilis str. 168 (bacteria)
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MGSSHHHHHH SSGLVPRGSH MLEFSVLKKL GWFFKAYWLR YTIAIVLLLA VNVIEMFPPK LLGNAIDDMK AGAFTAEGLL FYIGIFFVL TAAVYIMSYF WMHQLFGGAN LMEKILRTKL MGHLLTMSPP FYEKNRTGDL MARGTNDLQA VSLTTGFGIL T LVDSTMFM ...String:
MGSSHHHHHH SSGLVPRGSH MLEFSVLKKL GWFFKAYWLR YTIAIVLLLA VNVIEMFPPK LLGNAIDDMK AGAFTAEGLL FYIGIFFVL TAAVYIMSYF WMHQLFGGAN LMEKILRTKL MGHLLTMSPP FYEKNRTGDL MARGTNDLQA VSLTTGFGIL T LVDSTMFM MTIFLTMGFL ISWKLTFAAI IPLPVMAIAI SLYGSKIHER FTEAQNAFGA LNDRVLESVS GVRVIRAYVQ ET NDVRRFN EMTADVYQKN MKVAFIDSLF EPTVKLLVGA SYLIGLGYGA FLVFRNELTL GELVSFNVYL GMMIWPMFAI GEL INVMQR GNASLDRVNE TLSYETDVTD PKQPADLKEP GDIVFSHVSF TYPSSTSDNL QDISFTVRKG QTVGIAGKTG SGKT TIIKQ LLRQYPPGEG SITFSGVPIQ QIPLDRLRGW IGYVPQDHLL FSRTVKENIL YGKQDATDKE VQQAIAEAHF EKDLH MLPS GLETMVGEKG VALSGGQKQR ISIARALMAN PEILILDDSL SAVDAKTEAA IIKNIRENRK GKTTFILTHR LSAVEH ADL ILVMDGGVIA ERGTHQELLA NNGWYREQYE RQQLFTAEEG GAGA

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Macromolecule #2: Probable multidrug resistance ABC transporter ATP-binding/permeas...

MacromoleculeName: Probable multidrug resistance ABC transporter ATP-binding/permease protein YheH
type: protein_or_peptide / ID: 2 / Enantiomer: LEVO
EC number: Translocases; Catalysing the translocation of other compounds; Linked to the hydrolysis of a nucleoside triphosphate
Source (natural)Organism: Bacillus subtilis subsp. subtilis str. 168 (bacteria)
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MKIGKTLWRY ALLYRKLLIT AVLLLTVAVG AELTGPFIGK KMIDDHILGI EKTWYEAAEK DKNAVQFHGV SYVREDRLQE PVSKAKEAH IYQVGMAFYF VDQAVSFDGN RTVSDGKLTI TNGDKSRAYA AEKLTKQELF QFYQPEIKGM VLLICLYGGL L VFSVFFQY ...String:
MKIGKTLWRY ALLYRKLLIT AVLLLTVAVG AELTGPFIGK KMIDDHILGI EKTWYEAAEK DKNAVQFHGV SYVREDRLQE PVSKAKEAH IYQVGMAFYF VDQAVSFDGN RTVSDGKLTI TNGDKSRAYA AEKLTKQELF QFYQPEIKGM VLLICLYGGL L VFSVFFQY GQHYLLQMSA NRIIQKMRQD VFSHIQKMPI RYFDNLPAGK VVARITNDTE AIRDLYVTVL STFVTSGIYM FG IFTALFL LDVKLAFVCL AIVPIIWLWS VIYRRYASYY NQKIRSINSD INAKMNESIQ GMTIIQAFRH QKETMREFEE LNE SHFYFQ NRMLNLNSLM SHNLVNVIRN LAFVCLIWHF GGASLNAAGI VSIGVLYAFV DYLNRLFQPI TGIVNQFSKL ELAR VSAGR VFELLEEKNT EEAGEPAKER ALGRVEFRDV SFAYQEGEEV LKHISFTAQK GETVALVGHT GSGKSSILNL LFRFY DAQK GDVLIDGKSI YNMSRQELRS HMGIVLQDPY LFSGTIGSNV SLDDERMTEE EIKNALRQVG AEPLLKKLPK GINEPV IEK GSTLSSGERQ LISFARALAF DPAILILDEA TAHIDTETEA VIQKALDVVK QGRTTFVIAH RLSTIRNADQ ILVLDKG EI VERGNHEELM ALEGQYYQMY ELQKGQKHSI ALEHHHHHH

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration3 mg/mL
BufferpH: 8
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 15.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.2 µm / Nominal defocus min: 0.4 µm
Sample stageCooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 5.65 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 262341
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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