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Methods Mol Biol. 2019;1987:23-37. doi: 10.1007/978-1-4939-9446-5_2.

Expression, Purification, and Crystallization of the Transient Receptor Potential Channel TRPV6.

Author information

1
Department of Biochemistry and Molecular Biophysics, Columbia University, New York, NY, USA.
2
Integrated Program in Cellular, Molecular and Biomedical Studies, Columbia University, New York, NY, USA.
3
Department of Biochemistry and Molecular Biophysics, Columbia University, New York, NY, USA. as4005@cumc.columbia.edu.

Abstract

Transient receptor potential (TRP) channels are polymodal sensory transducers that respond to chemicals, temperature, mechanical stress, and membrane voltage and are involved in vision, taste, olfaction, hearing, touch, thermal perception, and nociception. TRP channels are implicated in numerous devastating diseases, including various forms of cancer, and represent important drug targets. The large sizes, low expression levels, and conformational dynamics of TRP channels make them challenging targets for structural biology. Here, we present the methodology used in structural studies of TRPV6, a TRP channel that is highly selective for calcium and mediates Ca2+ uptake in epithelial tissues. We provide a protocol for the expression, purification, and crystallization of TRPV6. Similar approaches can be used to determine crystal structures of other membrane proteins, including different members of the TRP channel family.

KEYWORDS:

Baculovirus; HEK 293 cells; Ion channel; Membrane protein; Protein crystallization; Protein expression; Protein purification; Protein structure; Sf9 cells; TRP channel; Transient receptor potential channel; X-ray crystallography

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