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- PDB-8q7b: ABCG2 in complex with MZ29 and 5D3 Fab -

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Basic information

Entry
Database: PDB / ID: 8q7b
TitleABCG2 in complex with MZ29 and 5D3 Fab
Components
  • 5D3(Fab) heavy chain variable domain
  • 5D3(Fab) light chain variable domain
  • ATP-binding cassette sub-family G member 2
KeywordsTRANSPORT PROTEIN / ABC transporter / multidrug resistance
Function / homology
Function and homology information


biotin transmembrane transporter activity / biotin transport / riboflavin transport / riboflavin transmembrane transporter activity / sphingolipid transporter activity / renal urate salt excretion / Abacavir transmembrane transport / urate metabolic process / sphingolipid biosynthetic process / urate transmembrane transporter activity ...biotin transmembrane transporter activity / biotin transport / riboflavin transport / riboflavin transmembrane transporter activity / sphingolipid transporter activity / renal urate salt excretion / Abacavir transmembrane transport / urate metabolic process / sphingolipid biosynthetic process / urate transmembrane transporter activity / Sphingolipid de novo biosynthesis / external side of apical plasma membrane / xenobiotic transport across blood-brain barrier / organic anion transport / : / transepithelial transport / Ciprofloxacin ADME / export across plasma membrane / Paracetamol ADME / NFE2L2 regulating MDR associated enzymes / Differentiation of Keratinocytes in Interfollicular Epidermis in Mammalian Skin / ABC-type xenobiotic transporter / Heme biosynthesis / cellular detoxification / ABC-type xenobiotic transporter activity / Heme degradation / efflux transmembrane transporter activity / ATPase-coupled transmembrane transporter activity / xenobiotic transmembrane transporter activity / transport across blood-brain barrier / brush border membrane / Iron uptake and transport / transmembrane transport / mitochondrial membrane / apical plasma membrane / membrane raft / protein homodimerization activity / ATP hydrolysis activity / nucleoplasm / ATP binding / identical protein binding / plasma membrane
Similarity search - Function
ABC transporter family G domain / ABC-2 type transporter / : / ABC-2 type transporter / ABC-2 type transporter / 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
Chem-BWQ / CHOLESTEROL / Broad substrate specificity ATP-binding cassette transporter ABCG2
Similarity search - Component
Biological speciesMus musculus (house mouse)
Homo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.56 Å
AuthorsKowal, J. / Yu, Q. / Ni, D. / Stahlberg, H. / Tajkhorshid, E. / Altmann, K.H. / Locher, K.P. / Manolaridis, I. / Jackson, S.M. / Taylor, N.M.I. / Zechner, M.
Funding support Switzerland, 1items
OrganizationGrant numberCountry
Swiss National Science FoundationTransCure Switzerland
Citation
Journal: ACS Chem Biol / Year: 2024
Title: Modulation of ABCG2 Transporter Activity by Ko143 Derivatives.
Authors: Qin Yu / Sepehr Dehghani-Ghahnaviyeh / Ali Rasouli / Anna Sadurni / Julia Kowal / Rose Bang-Soerensen / Po-Chao Wen / Melanie Tinzl-Zechner / Rossitza N Irobalieva / Dongchun Ni / Henning ...Authors: Qin Yu / Sepehr Dehghani-Ghahnaviyeh / Ali Rasouli / Anna Sadurni / Julia Kowal / Rose Bang-Soerensen / Po-Chao Wen / Melanie Tinzl-Zechner / Rossitza N Irobalieva / Dongchun Ni / Henning Stahlberg / Karl-Heinz Altmann / Emad Tajkhorshid / Kaspar P Locher /
Abstract: ABCG2 is a multidrug transporter that protects tissues from xenobiotics, affects drug pharmacokinetics, and contributes to multidrug resistance of cancer cells. Here, we present tetracyclic ...ABCG2 is a multidrug transporter that protects tissues from xenobiotics, affects drug pharmacokinetics, and contributes to multidrug resistance of cancer cells. Here, we present tetracyclic fumitremorgin C analog Ko143 derivatives, evaluate their modulation of purified ABCG2, and report four high-resolution cryo-EM structures and computational analyses to elucidate their interactions with ABCG2. We found that Ko143 derivatives that are based on a ring-opened scaffold no longer inhibit ABCG2-mediated transport activity. In contrast, closed-ring, tetracyclic analogs were highly potent inhibitors. Strikingly, the least potent of these compounds, MZ82, bound deeper into the central ABCG2 cavity than the other inhibitors and it led to partial closure of the transmembrane domains and increased flexibility of the nucleotide-binding domains. Minor structural modifications can thus convert a potent inhibitor into a compound that induces conformational changes in ABCG2 similar to those observed during binding of a substrate. Molecular dynamics simulations and free energy binding calculations further supported the correlation between reduced potency and distinct binding pose of the compounds. We introduce the highly potent inhibitor AZ99 that may exhibit improved stability.
#1: Journal: Nat Struct Mol Biol / Year: 2018
Title: Structural basis of small-molecule inhibition of human multidrug transporter ABCG2.
Authors: Scott M Jackson / Ioannis Manolaridis / Julia Kowal / Melanie Zechner / Nicholas M I Taylor / Manuel Bause / Stefanie Bauer / Ruben Bartholomaeus / Guenther Bernhardt / Burkhard Koenig / ...Authors: Scott M Jackson / Ioannis Manolaridis / Julia Kowal / Melanie Zechner / Nicholas M I Taylor / Manuel Bause / Stefanie Bauer / Ruben Bartholomaeus / Guenther Bernhardt / Burkhard Koenig / Armin Buschauer / Henning Stahlberg / Karl-Heinz Altmann / Kaspar P Locher /
Abstract: ABCG2 is an ATP-binding cassette (ABC) transporter that protects tissues against xenobiotics, affects the pharmacokinetics of drugs and contributes to multidrug resistance. Although many inhibitors ...ABCG2 is an ATP-binding cassette (ABC) transporter that protects tissues against xenobiotics, affects the pharmacokinetics of drugs and contributes to multidrug resistance. Although many inhibitors and modulators of ABCG2 have been developed, understanding their structure-activity relationship requires high-resolution structural insight. Here, we present cryo-EM structures of human ABCG2 bound to synthetic derivatives of the fumitremorgin C-related inhibitor Ko143 or the multidrug resistance modulator tariquidar. Both compounds are bound to the central, inward-facing cavity of ABCG2, blocking access for substrates and preventing conformational changes required for ATP hydrolysis. The high resolutions allowed for de novo building of the entire transporter and also revealed tightly bound phospholipids and cholesterol interacting with the lipid-exposed surface of the transmembrane domains (TMDs). Extensive chemical modifications of the Ko143 scaffold combined with in vitro functional analyses revealed the details of ABCG2 interactions with this compound family and provide a basis for the design of novel inhibitors and modulators.
History
DepositionAug 16, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Nov 6, 2024Provider: repository / Type: Initial release
Revision 1.1Nov 27, 2024Group: Data collection / Database references / Category: citation / citation_author / em_admin
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation_author.identifier_ORCID / _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
D: 5D3(Fab) heavy chain variable domain
E: 5D3(Fab) light chain variable domain
F: 5D3(Fab) heavy chain variable domain
B: ATP-binding cassette sub-family G member 2
A: ATP-binding cassette sub-family G member 2
C: 5D3(Fab) light chain variable domain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)245,41020
Polymers239,6476
Non-polymers5,76314
Water86548
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

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Antibody , 2 types, 4 molecules DFEC

#1: Antibody 5D3(Fab) heavy chain variable domain


Mass: 23843.633 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mus musculus (house mouse) / Production host: Escherichia coli (E. coli)
#2: Antibody 5D3(Fab) light chain variable domain


Mass: 23594.016 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mus musculus (house mouse) / Production host: Escherichia coli (E. coli)

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Protein / Sugars , 2 types, 4 molecules BA

#3: Protein ATP-binding cassette sub-family G member 2 / Breast cancer resistance protein / CDw338 / Mitoxantrone resistance-associated protein / Placenta- ...Breast cancer resistance protein / CDw338 / Mitoxantrone resistance-associated protein / Placenta-specific ATP-binding cassette transporter / Urate exporter


Mass: 72385.852 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: ABCG2, ABCP, BCRP, BCRP1, MXR / Production host: Homo sapiens (human)
References: UniProt: Q9UNQ0, ABC-type xenobiotic transporter
#4: Polysaccharide 2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 424.401 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DGlcpNAcb1-4DGlcpNAcb1-Glycam Condensed SequenceGMML 1.0
WURCS=2.0/1,2,1/[a2122h-1b_1-5_2*NCC/3=O]/1-1/a4-b1WURCSPDB2Glycan 1.1.0
[]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{}}}LINUCSPDB-CARE

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Non-polymers , 3 types, 60 molecules

#5: Chemical
ChemComp-CLR / CHOLESTEROL


Mass: 386.654 Da / Num. of mol.: 10 / Source method: obtained synthetically / Formula: C27H46O
#6: Chemical ChemComp-BWQ / ~{tert}-butyl 3-[(2~{S},5~{S},8~{S})-14-cyclopentyloxy-2-(2-methylpropyl)-4,7-bis(oxidanylidene)-3,6,17-triazatetracyclo[8.7.0.0^{3,8}.0^{11,16}]heptadeca-1(10),11,13,15-tetraen-5-yl]propanoate


Mass: 523.664 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C30H41N3O5 / Feature type: SUBJECT OF INVESTIGATION
#7: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 48 / Source method: isolated from a natural source / Formula: H2O

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Details

Has ligand of interestY
Has protein modificationY

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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: ABCG2 in complex with MZ29 and 5D3 Fab / Type: COMPLEX
Details: Nanodisc-reconstituted 5D3-Fab-ABCG2 was incubated with MZ29
Entity ID: #1-#3 / Source: RECOMBINANT
Molecular weightValue: 0.244 MDa / Experimental value: YES
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Homo sapiens (human) / Cell: HEK293EBNA
Buffer solutionpH: 7.5
Buffer componentConc.: 0.04 M / Name: Hepes
SpecimenConc.: 0.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES / Details: The sample was mono-disperse.
Specimen supportGrid material: COPPER / Grid mesh size: 400 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE-PROPANE / 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 FIELD / Nominal magnification: 130000 X / Nominal defocus max: 2200 nm / Nominal defocus min: 800 nm / Cs: 2.7 mm / C2 aperture diameter: 100 µm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 99 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K2 SUMMIT (4k x 4k)
EM imaging opticsEnergyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV

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Processing

EM software
IDNameVersionCategory
2EPU2image acquisition
7UCSF Chimeramodel fitting
9RELION3.1initial Euler assignment
10RELION3.1final Euler assignment
11RELION3.1classification
12RELION3.13D reconstruction
13PHENIXmodel refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C2 (2 fold cyclic)
3D reconstructionResolution: 2.56 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 205986 / Symmetry type: POINT
Atomic model buildingB value: 50
Atomic model buildingPDB-ID: 6ETI

6eti


Accession code: 6ETI / Source name: PDB / Type: experimental model
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00713034
ELECTRON MICROSCOPYf_angle_d1.61217734
ELECTRON MICROSCOPYf_dihedral_angle_d16.0254650
ELECTRON MICROSCOPYf_chiral_restr0.1972052
ELECTRON MICROSCOPYf_plane_restr0.0052144

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