苜蓿盲蝽气味分子结合蛋白AlinOBP2结合特性初步分析

Preliminary Binding Characterization of Odorant Binding Protein 2 in the Lucerne Plant Bug, Adelphocoris lineolatus (Goeze)

昆虫具有灵敏的嗅觉系统,能够特异性地识别性信息素和寄主挥发物来进行寻找配偶、定位寄主植物和产卵位点.气味分子结合蛋白在昆虫嗅觉识别过程中发挥关键作用.本研究表达和纯化了一个新的苜蓿盲蝽气味分子结合蛋白AlinOBP2,采用qRT-PCR方法解析了AlinOBP2基因的表达谱,结果表明AlinOBP2绝大部分在触角中表达,且在雌雄触角中的表达量相当,在头部也有少量的表达.以N-phenyl-1-naphthylamine(1-NPN)为荧光探针,采用荧光竞争结合实验研究了5种性信息素类似物和13种棉花挥发物与AlinOBP2蛋白的结合能力.结果显示,5种性信息素类似物均不能和AlinOBP2有效结合,暗示AlinOBP2在苜蓿盲蝽寻找配偶过程中不发挥作用.在13种棉花挥发物中,庚酸乙酯和AlinOBP2的结合能力最强,结合常数为9.22μmol/L.二甲基萘、3-己酮、乙酸叶醇酯、乙酸壬酯、香芹醇5种化合物和AlinOBP2结合能力一般,结合常数分别为15.49,17.31,21.53,18.86和13.47μmol/L.据此推测,AlinOBP2可能为普通气味结合蛋白,能够选择性地结合某些棉花挥发物并参与苜蓿盲蝽识别普通气味过程.

Insects can use their sensitive olfactory system to perceive sex pheromones and host volatiles, and trigger specific behavioral reactions, such as detection and location mate partners, host plants and oviposition sites. Odorant binding proteins （OBPs） have been demonstrated to play a crucial role in the olfaction perception in insects. In the present study, a novel OBP protein （AlinOBP2） in the lucerne plant bug, Adelphocoris lineolatus, was expressed and purified. The expression intensity of AlinOBP2 gene was evaluated by qRT-PCR, which indicated that AlinOBP2 was mainly expressed in antennae. The expression level is nearly equivalent between the adult male and female antennae. In addition, AlinOBP2 was also expressed in the head with a much lower expression intensity. The binding properties of AlinOBP2 with five sex pheromone analogs and 13 cotton volatiles were measured by fluorescence competitive binding assays with the fluorescence probe N-phenyl-l-naphthylamine （1-NPN）. The results reveal that AlinOBP2 cannot effectively bind with the five sex pheromone analogs, indicating AlinOBP2 did not participate in the sex pheromone reception process. Of the 13 cotton volatiles, ethyl heptanoate had the strongest binding affinity with AlinOBP2, with dissociation constant as 9.22 μmol/L. 2-methylnaphthalene, 3-hexanone, cis-3-hexenyl acetate, nonyl acetate, and carveol had medium binding affinities with AlinOBP2, with dissociation constants of 15.49, 17.31, 21.53, 18.86 and 13.47 μmol/L, respectively. The results suggest that AlinOBP2 is a general odorant binding protein （GOBP） and can selectively bind with particular cotton volatiles and participate in the general odorant reception process.