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Updated: Nov 16, 2025

Method for Efficient Refolding and Purification of Chemoreceptor Ligand Binding Domain
Published on: December 12, 2017
Refolding and characterization of two G protein-coupled receptors purified from E. coli inclusion bodies
Bastian Heim1, René Handrick1, Marcus D Hartmann2
1Institute of Applied Biotechnology, University of Applied Sciences, Biberach, Germany.
Abstract:
Aiming at streamlining GPCR production from E. coli inclusion bodies for structural analysis, we present a generic approach to assess and optimize refolding yield through thermostability analysis. Since commonly used hydrophobic dyes cannot be applied as probes for membrane protein unfolding, we adapted a technique based on reacting cysteins exposed upon thermal denaturation with fluorescent 7-Diethylamino-3-(4-maleimidophenyl)-4-methylcoumarin (CPM). Successful expression, purification and refolding is shown for two G protein-coupled receptors (GPCR), the sphingosine-1-phosphate receptor S1P1, and the orphan receptor GPR3. Refolded receptors were subjected to lipidic cubic phase crystallization screening.
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