Purification and 3D Structural Analysis of Oligomeric Human Multidrug Transporter ABCG2

Christopher A. McDevitt, Richard F. Collins, Michael Conway, S. Modok, Janet Storm, Ian D. Kerr, Robert C. Ford, Richard Callaghan

Research output: Contribution to journalArticle

97 Citations (Scopus)

Abstract

ABCG2 is a multidrug efflux pump associated with resistance of cancer cells to a plethora of unrelated drugs. ABCG2 is a "half-transporter," and previous studies have indicated that it forms homodimers and higher oligomeric species. In this manuscript, electron microscopic structural analysis directly addressed this issue. An N-terminal hexahistidine-tagged ABCG2R482G isoform was expressed to high levels in insect cells. An extensive detergent screen was employed to effect extraction of ABCG2R482G from membranes and identified only the fos-choline detergents as efficient. Soluble protein was purified to >95% homogeneity by a three-step procedure while retaining the ability to bind substrates. Cryonegative stain electron microscopy of purified ABCG2R482G provided 3D structural data at a resolution of ∼18 Å. Single-particle analysis revealed that the complex forms a tetrameric complex (∼180 Å in diameter × ∼140 Å high) with an aqueous central region. We interpret the tetrameric structure as comprising four homodimeric ABCG2R482G complexes.

Original languageEnglish
Pages (from-to)1623-1632
Number of pages10
JournalStructure
Volume14
Issue number11
DOIs
Publication statusPublished - Nov 2006

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His-His-His-His-His-His
Detergents
Choline
Insects
Electron Microscopy
Protein Isoforms
Coloring Agents
Electrons
Membranes
Pharmaceutical Preparations
Neoplasms
Proteins
ATP Binding Cassette Transporter, Sub-Family G, Member 2

Keywords

  • CELLBIO
  • CELLCYCLE

ASJC Scopus subject areas

  • Molecular Biology
  • Structural Biology

Cite this

McDevitt, C. A., Collins, R. F., Conway, M., Modok, S., Storm, J., Kerr, I. D., ... Callaghan, R. (2006). Purification and 3D Structural Analysis of Oligomeric Human Multidrug Transporter ABCG2. Structure, 14(11), 1623-1632. https://doi.org/10.1016/j.str.2006.08.014

Purification and 3D Structural Analysis of Oligomeric Human Multidrug Transporter ABCG2. / McDevitt, Christopher A.; Collins, Richard F.; Conway, Michael; Modok, S.; Storm, Janet; Kerr, Ian D.; Ford, Robert C.; Callaghan, Richard.

In: Structure, Vol. 14, No. 11, 11.2006, p. 1623-1632.

Research output: Contribution to journalArticle

McDevitt, CA, Collins, RF, Conway, M, Modok, S, Storm, J, Kerr, ID, Ford, RC & Callaghan, R 2006, 'Purification and 3D Structural Analysis of Oligomeric Human Multidrug Transporter ABCG2', Structure, vol. 14, no. 11, pp. 1623-1632. https://doi.org/10.1016/j.str.2006.08.014
McDevitt, Christopher A. ; Collins, Richard F. ; Conway, Michael ; Modok, S. ; Storm, Janet ; Kerr, Ian D. ; Ford, Robert C. ; Callaghan, Richard. / Purification and 3D Structural Analysis of Oligomeric Human Multidrug Transporter ABCG2. In: Structure. 2006 ; Vol. 14, No. 11. pp. 1623-1632.
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