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触角结合蛋白(antennal binding proteins,ABPs)是气味结合蛋白(odorant binding proteins,OBPs)的一个亚类,推测其在昆虫嗅觉中起作用。为了探讨这一问题,本研究通过转录组数据分析并利用RACE技术,克隆了甜菜夜蛾Spodoptera exigua触角结合蛋白Ⅱ基因(SexigABP2)的全长cDNA序列(GenBank登录号为HQ234486)。序列分析表明,该基因开放阅读框长444bp,编码148个氨基酸,具有OBPs典型的6个半胱氨酸位点;其氨基酸序列和烟芽夜蛾Heliothisvirescens的HvirABP2的一致性最高,达72%。实时定量PCR分析显示,该基因主要在触角中表达,在喙、足、翅等组织中也有少量表达,且在雌蛾触角及足中的表达量显著高于雄蛾。进一步对该基因进行原核表达和纯化,利用荧光竞争结合实验测定了SexigABP2对35种气味物质的结合能力,发现其对甜菜夜蛾性信息素组分(Z)-9-十四碳烯醇和植物挥发物法尼醇的结合能力较强,结合常数分别为8.24μmol/L和8.14μmol/L。结合能力比较表明,SexigABP2对不饱和长碳链化合物较饱和短碳链化合物具有更强的结合能力;在不饱和长碳链化合物中,对醇类物质又较乙酸酯类物质具有更强的结合能力。结果提示SexigABP2可能参与了成虫对不饱和长碳链的植物挥发物的感受。
The antennal binding proteins (ABPs), a subset of odorant binding proteins (OBPs), are presumed to play an important role in insect olfactory. In order to explore this issue, the full-length cDNA sequence of Spodoptera exigua angiotensin Ⅱ binding protein Ⅱ (GenBank accession number HQ234486) was cloned by transcriptome analysis and RACE technology. Sequence analysis showed that the open reading frame of this gene was 444bp in length and encoded 148 amino acids with 6 typical cysteine residues of OBPs. The amino acid sequence of the gene was identical with HvirABP2 of Heliothis virescens by 72%. Real-time quantitative PCR analysis showed that the gene was mainly expressed in the antennae, but also in the beak, the foot, the wing and other tissues, and the expression level in the antennae and the foot of the female was significantly higher than that in the male. The prokaryotic expression and purification of this gene was further studied. The binding ability of SexigABP2 to 35 odorant compounds was determined by fluorescence-mediated competition assay. The results showed that the binding of SexigABP2 to sex pheromone component (Z) -9-tetradecenol and plant Volatile alcohol farnesyl binding ability, binding constants were 8.24μmol / L and 8.14μmol / L. The binding capacity shows that SexigABP2 has stronger binding ability to unsaturated long-chain compounds than saturated short-chain compounds. In the unsaturated long-chain compounds, the alcohol compounds have stronger binding to acetic acid esters ability. The results suggest that SexigABP2 may be involved in the perception of plant volatiles on unsaturated long carbon chains.