| Apolygus lucorum is an omnivorous field pest with a wide host range.It can harm cotton,mung bean,jujube trees and tea trees in the field which causes serious economic losses.In recent years,the research of the green prevention and control of A.lucorum quickly with the rapid development of food and sex attractants which is based on the volatile components of the host and the sex pheromones of A.lucorum,and upgraded traps.In the chemosensory process of insects,odorant binding proteins(OBPs)can bind and transfer of small chemical messages in the environment that cause the behavioral responses of insects,such as host switching,feeding,and mating.Therefore,it is of great significance to study the binding characteristics and functions of OBPs for the development of efficient and green prevention and control technology of A.lucorum.In this study,molecular biology and Electroantennogram(EAG)technology were used to reveal the binding characteristics of 12 AlucOBPs from the A.lucorum and to speculate on their potential functions at the protein and physiological levels.At the same time,combined with the existing research on AlucOBPs,we summarized the the binding characteristics and common capacities of 38 AlucOBPs.It is helpful to clarify the mechanism of olfactory recognition of A.lucorum on host volatiles and sex pheromones,filter out potential target sites and novel behavior control compounds,and provide theoretical advice for formulating green and effective strategies for the prevention and treatment of A.lucorum.The main results are as follows:1.Induced expression and purification of 12 AlucOBPs in A.lucorumIn this study,12 AlucOBPs were induced by recombinant expression technique in vitro,and purified by nickel column affinity chromatography.The results showed that 12 AlucOBPs were successfully induced and purified,of which 5 AlucOBPs were expressed in the supernatant of the bacterial solution and 7 AlucOBPs were expressed in the inclusion body of the bacterial solution.2.Binding characteristics and potential function prediction of 12 AlucOBPs in A.lucorumThe binding characteristics of 12 AlucOBPs,which have not been fully studied.were determined by fluorescence competitive binding test,and their functions were analyzed.The results showed that all the 8 AlucOBPs could bind to host volatiles and other substances.suggesting that these proteins had multiple functions.AlucOBP18 and AlucOBP31 not only bind to host volatiles and other substances,but also bind to the sex pheromone of the species.which is presumed to be sex pheromone bindingotther proteins,indicating that these two proteins play an significant role in mating and oviposition of the species.Secondly,AlucOBP4,AlucOBP16,AlucOBP20 and AlucOBP23 only bind to taste substances.indicating that these four odor binding proteins are mainly involved in the taste recognition process of A.lucorum and have taste function.3.EAG reaction between 12 volatile binding ligands of AlucOBP31 and antennae of A.lucorumThe EAG reactions of 12 volatile binding ligands of AlucOBP31 with A.lucorum were detected by antennae potentiometric technique.The results showed that all the 12 volatiles(100 mg/mL)induced potential responses in the antennae of A.lucorum,but there were some differences among the reactions.Among them,the relative potential response values of the antennae to(E)-4-oxo-2-hexenal and cis-3-Hexenyl butyrate were the highest,and the male antennae were significantly higher than the female antennae(P<0.05).In addition.the potential responses of Decanal,DMNT,Dodecyl aldehyde and Ocimene to antennae were also significantly different between males and females(P<0.05).At the same time,(E)-4oxo-2-hexenal and cis-3-Hexenyl butyrate had dose-effect on the relative reaction value of EAG in the antennae of A.lucorum,and the relative reaction value of EAG increased with the increase of compound concentration.4.Binding characteristics and functional analysis of 38 AlucOBPs from A.lucorumCombined with previous studies on AlucOBPs,a binding map was made to summarize the binding characteristics and functional characteristics of 38 AlucOBPs.The results showed that the binding spectra of AlucOBP1,AlucOBP3,AlucOBP 18 and AlucOBP 31 were wider.At the same time,most of the proteins in the 38 AlucOBPs could bind to βionone and β-ionone derivatives,and the binding ability was strong.According to functional analysis,the 38 AlucOBPs included 12 PBPs and 26 GOBPs,and most of the PBPs were distributed equally in the head and antennae ofA.lucorum.In addition.most AlucOBPs have multiple functions. |
