Human N-formyl peptide receptors (FPRs) belong to the G protein-coupled receptors (GPCRs) superfamily, the
most frequently addressed drug targets in the pharmaceutical industry, and are considered to play important roles in innate
immunity and host defense mechanisms. Although still a highly challenging task, the availability of soluble and functional
GPCRs including FPRs in milligram quantities is essential to spur the advancement of protein-based structural and functional
studies for drug discovery. In this report, the applicability of E. coli extracts-based cell-free expression system to
producing soluble and active human FPRs and hence to FPRs protein-based research was evaluated, during which human
FPR3 was selected as our prototype receptor. To better solubilize the freshly expressed human FPR3, a panel of different
detergents (mostly nonionic detergents) were selected and evaluated in the cell-free system devoid of natural membrane.
After one-step immunoaffinity purification, the secondary structure and biological function of purified FPR3 were characterized.
A final yield of 0.6 mg functional human FPR3 per ml reaction volume was obtained. The demonstrated proper
folding and functionality of the cell-free produced human FPR3 opens a new avenue for the fast and efficient generation
of human FPRs (and even other GPCRs) for structural and functional analysis.
Keywords: Biological function, cell-free expression system, detergent, G protein-coupled receptors, N-formyl peptide receptors,
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