Abstract
This chapter outlines a protocol developed to prepare a purified deuterated membrane protein for a small-angle neutron scattering (SANS) experiment. SANS is a noninvasive technique well suited to studying membrane protein solution structures, and deuteration enhances the signal from the protein over the background (Breyton et al., Eur Phys J E Soft Matter 36 (7):71, 2013; Garg et al., Biophys J 101 (2):370–377, 2011). We present our workflow: transformation of our plasmid into E. coli, cell growth and expression of our deuterated protein, membrane isolation, detergent solubilization, protein purification, purity assessment, and final preparation for SANS.
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Acknowledgments
SANS studies on IAPs in the Lieberman lab are supported by NSF grant 1817796. The Office of Biological and Environmental Research supported research at Oak Ridge National Laboratory’s Center for Structural Molecular Biology (CSMB), using facilities supported by the Scientific User Facilities Division, Office of Basic Energy Sciences, US Department of Energy.
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Wu, Y., Weiss, K.L., Lieberman, R.L. (2021). Preparation of a Deuterated Membrane Protein for Small-Angle Neutron Scattering. In: Schmidt-Krey, I., Gumbart, J.C. (eds) Structure and Function of Membrane Proteins. Methods in Molecular Biology, vol 2302. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1394-8_12
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