Functional Analysis of PxylPBP2 Responding to Repellent Activity of Natural Pyrazine Against Diamondback Moth.

2,3-dimethyl-6-(1-hydroxy)-pyrazine showed significant repellent activity to diamondback moth (Plutella xylostella), the PxylPBP2 was considered as the potential molecular target, though this has not been validated. In this study, the repellent rate was significantly decreased by about 20% when the PxylPBP2 was silenced using RNAi. There was obvious interaction between 2,3-dimethyl-6-(1-hydroxy)-pyrazine and PxylPBP2, with a Kd (Dissociation constant) value of 7.78 μmol/L via microscale thermophoresis, while the value was changed to 4.45 μmol/L when the PxylOR31 (Pxyl, Plutella xylostella; OR31, Odorant receptor 31) was introduced to this compound and ligand system. Moreover, the RMSD (Root mean square deviation) value of pyrazine + PxylPBP2 + PxylOR31 treatment ranged between 0.030 and 0.125 nm; the average number of hydrogen bonds and Van der Waals interaction value can be used to summarize that there was a stronger interaction between pyrazine and PxylPBP2. Furthermore, the ILE122 (L-Isoleucine at 122 site) was an important target site during the binding process between PxylBPB2 and ligands, which was validated by performing a fluorescence competitive binding experiment. Finally, the PxylPBP2 showed a strong ability to bind to α-pinene and β-ionone with the Ki values of 10.54 μmol/L and 11.74 μmol/L, respectively. These data provide a creative strategy for developing novel insecticides, which is also important for achieving the green control of P. xylostella and delaying the development of resistance.