Methods Enzymol 2020 Jan 01;642:279-300. doi: 10.1016/bs.mie.2020.04.065.

Effect of OBPs on the response of olfactory receptors.

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The technique of two-electrode voltage-clamp (TEVC) recording from the heterologous expression system of olfactory receptors (ORs) in Xenopus laevis oocytes has been widely used to deorphanize insect ORs, that is to identify specific ligands for each of them. However, there is a controversial issue on whether ORs are activated by the odorant/OBP complex or the odorant alone. The mechanism of interaction among odorants, odorant-binding proteins (OBPs) and ORs remains largely unknown, due to the limitations in the use of scientific and innovative methods. In this chapter, the modified Xenopus oocytes expression system combined with TEVC technique is used to approach this issue. We describe the experimental strategies and provide detailed protocols for recording the signals generated by ORs in response to odorant/OBP complex at different concentrations. Results obtained by this approach have revealed that the presence of OBPs in the system affects the selectivity and sensitivity responses of ORs. Such studies help understanding the molecular mechanism of odorant detection in peripheral nervous system.

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Species referenced: Xenopus laevis