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Nanodiscs in the Studies of Membrane-Bound Cytochrome P450 Enzymes

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Cytochrome P450 Protocols

Part of the book series: Methods in Molecular Biology ((MIMB,volume 987))

Abstract

Cytochromes P450 from eukaryotes and their native redox partners cytochrome P450 reductases both belong to the class of monotopic membrane proteins containing one transmembrane anchor. Incorporation into the lipid bilayer significantly affects their equilibrium and kinetic properties and plays an important role in their interactions. We describe here the detailed protocols developed in our group for the functional self-assembly of mammalian cytochromes P450 and cytochrome P450 reductases into Nanodiscs with controlled lipid composition. The resulting preparations are fully functional, homogeneous in size, composition and oligomerization state of the heme enzyme, and show an improved stability with respect to P420 formation. We provide a brief overview of applications of Nanodisc technology to the biophysical and biochemical mechanistic studies of cytochromes P450 involved in steroidogenesis, and of the most abundant xenobiotic-metabolizing human cytochrome P450 CYP3A4.

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Acknowledgments

We gratefully acknowledge the valuable contribution to development of these methods made by T. H. Bayburt, B. J. Baas, M. A. McLean, and other members of the Sligar lab, whose works are cited in the list of references. This work was supported by NIH grants GM31756 and GM33775 to S.G. Sligar.

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Authors and Affiliations

  1. Department of Biochemistry, University of Illinois, Urbana, IL, USA

    A. Luthra, M. Gregory, Y. V. Grinkova & I. G. Denisov

  2. Department of Biochemistry, Beckman Institute and School of Molecular and Cellular Biology, University of Illinois, Urbana, IL, USA

    S. G. Sligar

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  1. A. Luthra
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  2. M. Gregory
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  3. Y. V. Grinkova
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  4. I. G. Denisov
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  5. S. G. Sligar
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Editors and Affiliations

  1. Sch. of Biological & Chemical Sciences, Queen Mary, University of London, Mile End Road, London, E1 4NS, United Kingdom

    Ian R. Phillips

  2. , Department of Structural and Molecular B, University College London, Gower Street, London, WC1E 6BT, United Kingdom

    Elizabeth A. Shephard

  3. University of California, 16th St 600, San Francisco, 94158-2517, California, USA

    Paul R. Ortiz de Montellano

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Luthra, A., Gregory, M., Grinkova, Y.V., Denisov, I.G., Sligar, S.G. (2013). Nanodiscs in the Studies of Membrane-Bound Cytochrome P450 Enzymes. In: Phillips, I., Shephard, E., Ortiz de Montellano, P. (eds) Cytochrome P450 Protocols. Methods in Molecular Biology, vol 987. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-321-3_10

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  • DOI: https://doi.org/10.1007/978-1-62703-321-3_10

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