Molecular and Functional Characterization of Pheromone Binding Protein 2 from Cyrtotrachelus buqueti (Coleoptera: Curculionidae)

Pheromone-binding proteins (PBPs) play important roles in binding and transporting sex pheromones. However, thePBPgenes identified in coleopteran insects and their information sensing mechanism are largely unknown.Cyrtotrachelus buqueti(Coleoptera: Curculionidae) is a major insect pest of bamboo plantations. In this study, a novelPBPgene,CbuqPBP2, fromC. buquetiwas functionally characterized.CbuqPBP2was more abundantly expressed in the antennae of both sexes than other body parts, and its expression level was significantly male-biased. Fluorescence competitive binding assays showed that CbuqPBP2 exhibited the strongest binding affinity to dibutyl phthalate (Ki= 6.32 μM), followed by styrene (Ki= 11.37 μM), among twelveC. buquetivolatiles. CbuqPBP2, on the other hand, showed high binding affinity to linalool (Ki= 10.55), the main volatile of host plantNeosinocalamus affinis. Furthermore, molecular docking also demonstrated the strong binding ability of CbuqPBP2 to dibutyl phthalate, styrene, and linalool, with binding energy values of −5.7, −6.6, and −6.0 kcal/mol, respectively, and hydrophobic interactions were the prevailing forces. The knockdown ofCbuqPBP2expression via RNA interference significantly reduced the electroantennography (EAG) responses of male adults to dibutyl phthalate and styrene. In conclusion, these results will be conducive to understanding the olfactory mechanisms ofC. buquetiand promoting the development of novel strategies for controlling this insect pest.Keywords:Cyrtotrachelus buqueti;pheromone-binding proteins;binding assay;molecular docking;RNA interference;electroantennography