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- EMDB-11670: Nanodisc reconstituted human ABCB1 in complex with MRK16 Fab and ... -

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Basic information

Entry
Database: EMDB / ID: EMD-11670
TitleNanodisc reconstituted human ABCB1 in complex with MRK16 Fab and elacridar
Map data
SampleNanodisc reconstituted human ABCB1 in complex with MRK16 Fab and elacridar
  • (Multidrug resistance protein ...ATP-binding cassette transporter) x 2
  • (MRK16 Fab-fragment ...) x 2
  • If kappa light chain
  • (ligand) x 2
Function / homology
Function and homology information


ceramide translocation / ceramide floppase activity / positive regulation of anion channel activity / floppase activity / external side of apical plasma membrane / transporter activity / phosphatidylcholine floppase activity / phosphatidylethanolamine flippase activity / regulation of response to osmotic stress / xenobiotic transport across blood-brain barrier ...ceramide translocation / ceramide floppase activity / positive regulation of anion channel activity / floppase activity / external side of apical plasma membrane / transporter activity / phosphatidylcholine floppase activity / phosphatidylethanolamine flippase activity / regulation of response to osmotic stress / xenobiotic transport across blood-brain barrier / ABC-type xenobiotic transporter / P-type phospholipid transporter / export across plasma membrane / stem cell proliferation / transepithelial transport / ATPase-coupled xenobiotic transmembrane transporter activity / phospholipid translocation / xenobiotic transmembrane transporter activity / transport across blood-brain barrier / efflux transmembrane transporter activity / ATPase-coupled transmembrane transporter activity / regulation of chloride transport / transmembrane transport / G2/M transition of mitotic cell cycle / apical plasma membrane / response to drug / ATPase activity / ubiquitin protein ligase binding / cell surface / extracellular exosome / membrane / integral component of membrane / ATP binding / plasma membrane / metal ion binding
Immunoglobulin subtype / Immunoglobulin V-set domain / Immunoglobulin/major histocompatibility complex, conserved site / ABC transporter-like / AAA+ ATPase domain / Immunoglobulin C1-set / Immunoglobulin-like domain / ABC transporter type 1, transmembrane domain / Immunoglobulin-like fold / ABC transporter, conserved site ...Immunoglobulin subtype / Immunoglobulin V-set domain / Immunoglobulin/major histocompatibility complex, conserved site / ABC transporter-like / AAA+ ATPase domain / Immunoglobulin C1-set / Immunoglobulin-like domain / ABC transporter type 1, transmembrane domain / Immunoglobulin-like fold / ABC transporter, conserved site / P-loop containing nucleoside triphosphate hydrolase / Multidrug resistance protein 1 / Immunoglobulin-like domain superfamily / ABC transporter type 1, transmembrane domain superfamily / Type I protein exporter
If kappa light chain / ATP-dependent translocase ABCB1
Biological speciesHomo sapiens (human) / Mus musculus (house mouse)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.6 Å
AuthorsNosol K / Locher KP
Funding support Switzerland, 1 items
OrganizationGrant numberCountry
Swiss National Science Foundation310030_189111 Switzerland
CitationJournal: Proc Natl Acad Sci U S A / Year: 2020
Title: Cryo-EM structures reveal distinct mechanisms of inhibition of the human multidrug transporter ABCB1.
Authors: Kamil Nosol / Ksenija Romane / Rossitza N Irobalieva / Amer Alam / Julia Kowal / Naoya Fujita / Kaspar P Locher /
Abstract: ABCB1 detoxifies cells by exporting diverse xenobiotic compounds, thereby limiting drug disposition and contributing to multidrug resistance in cancer cells. Multiple small-molecule inhibitors and ...ABCB1 detoxifies cells by exporting diverse xenobiotic compounds, thereby limiting drug disposition and contributing to multidrug resistance in cancer cells. Multiple small-molecule inhibitors and inhibitory antibodies have been developed for therapeutic applications, but the structural basis of their activity is insufficiently understood. We determined cryo-EM structures of nanodisc-reconstituted, human ABCB1 in complex with the Fab fragment of the inhibitory, monoclonal antibody MRK16 and bound to a substrate (the antitumor drug vincristine) or to the potent inhibitors elacridar, tariquidar, or zosuquidar. We found that inhibitors bound in pairs, with one molecule lodged in the central drug-binding pocket and a second extending into a phenylalanine-rich cavity that we termed the "access tunnel." This finding explains how inhibitors can act as substrates at low concentration, but interfere with the early steps of the peristaltic extrusion mechanism at higher concentration. Our structural data will also help the development of more potent and selective ABCB1 inhibitors.
Validation ReportPDB-ID: 7a6c

SummaryFull reportAbout validation report
History
DepositionAug 25, 2020-
Header (metadata) releaseOct 14, 2020-
Map releaseOct 14, 2020-
UpdateOct 14, 2020-
Current statusOct 14, 2020Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.01
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.01
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7a6c
  • Surface level: 0.01
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-7a6c
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_11670.map.gz / Format: CCP4 / Size: 216 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.84 Å/pix.
x 384 pix.
= 322.56 Å
0.84 Å/pix.
x 384 pix.
= 322.56 Å
0.84 Å/pix.
x 384 pix.
= 322.56 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.84 Å
Density
Contour LevelBy AUTHOR: 0.01 / Movie #1: 0.01
Minimum - Maximum-0.034995176 - 0.061851177
Average (Standard dev.)0.000008239 (±0.0010864616)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions384384384
Spacing384384384
CellA=B=C: 322.56 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.840.840.84
M x/y/z384384384
origin x/y/z0.0000.0000.000
length x/y/z322.560322.560322.560
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ161186271
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS384384384
D min/max/mean-0.0350.0620.000

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Supplemental data

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Sample components

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Entire Nanodisc reconstituted human ABCB1 in complex with MRK16 Fab and ...

EntireName: Nanodisc reconstituted human ABCB1 in complex with MRK16 Fab and elacridar
Number of components: 8

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Component #1: protein, Nanodisc reconstituted human ABCB1 in complex with MRK16...

ProteinName: Nanodisc reconstituted human ABCB1 in complex with MRK16 Fab and elacridar
Recombinant expression: No
MassExperimental: 240 kDa

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Component #2: protein, Multidrug resistance protein 1

ProteinName: Multidrug resistance protein 1Multiple drug resistance
Recombinant expression: No
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Homo sapiens (human)

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Component #3: protein, MRK16 Fab-fragment light and heavy chain

ProteinName: MRK16 Fab-fragment light and heavy chain / Recombinant expression: No
SourceSpecies: Mus musculus (house mouse)
Source (engineered)Expression System: Homo sapiens (human)

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Component #4: protein, Multidrug resistance protein 1

ProteinName: Multidrug resistance protein 1Multiple drug resistance
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 141.628781 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Homo sapiens (human)

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Component #5: protein, If kappa light chain

ProteinName: If kappa light chain / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 24.139758 kDa
SourceSpecies: Mus musculus (house mouse)
Source (engineered)Expression System: Homo sapiens (human)

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Component #6: protein, MRK16 Fab-fragment heavy chain

ProteinName: MRK16 Fab-fragment heavy chain / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 23.415236 kDa
SourceSpecies: Mus musculus (house mouse)
Source (engineered)Expression System: Homo sapiens (human)

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Component #7: ligand, elacridar

LigandName: elacridar / Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 0.563643 kDa

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Component #8: ligand, CHOLESTEROL

LigandName: CHOLESTEROL / Number of Copies: 10 / Recombinant expression: No
MassTheoretical: 0.386654 kDa

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Experimental details

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Sample preparation

SpecimenSpecimen state: Particle / Method: cryo EM
Sample solutionSpecimen conc.: 0.4 mg/mL / pH: 7.4
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 32 e/Å2 / Illumination mode: FLOOD BEAM
LensCs: 2.7 mm / Imaging mode: BRIGHT FIELD
Specimen HolderModel: FEI TITAN KRIOS AUTOGRID HOLDER
CameraDetector: OTHER

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Image processing

ProcessingMethod: single particle reconstruction / Number of projections: 241885
3D reconstructionResolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF
FSC plot (resolution estimation)

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Atomic model buiding

Output model

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