摘要
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。
Antennal binding proteins （ABPs） represent a sub-class of odorant binding proteins （OBPs）, and thus are assumed to play a role in insect olfaction. In order to explore the role of ABPs in olfaction, the full-length cDNA of an antennal binding protein 2 gene from Spodoptera exigua （SexigABP2） （GenBank accession no. HQ234486 ） was identified by transcriptome analysis and RACE technology. The sequence analysis showed that SexigABP2 contains a 444 bp open reading frame that encodes 148 amino acids including the six conserved cysteine residues of typical OBPs. SexigABP2 shares the highest amino acid identity （up to 72% ） with an ABP2 from Heliothis virescens （HvirABP2）. The results of real- time quantitative PCR showed that SexigABP2 was highly expressed in male and female antennae, but weakly expressed in proboscis, legs, and wings of both sexes. The expression levels in female antennae and legs were significantly higher than those in male antennae and legs, respectively. SexigABP2 was further expressed in a prokaryotic expression system, and the protein was purified. By fluorescence competitive binding assay, the affinities of SexigABP2 with 35 odorant compounds were tested. Among the tested ligands, （Z）-9-tetradeeenol （a sex pheromone component of S. exigua） and farnesol （a plant volatile compound） showed the highest affinity, with the Ki values of 8.24 μmol/L and 8.14 μmol/L, respectively. Affinity comparison indicated that long carbon-chain compounds with unsaturated bond（s） exhibited the higher affinities than short ones without unsaturated bond; among the unsaturated long carbon-chain compounds, however, alcohols displayed higher affinities than acetates. The results suggest that SexigABP2 might be involved in perception of plant volatile compounds with a long carbon-chain and unsaturated bonds.
出处
《昆虫学报》
CAS
CSCD
北大核心
2012年第5期499-509,共11页
Acta Entomologica Sinica
基金
国家自然科学基金项目(31071978)
关键词
甜菜夜蛾
触角结合蛋白
基因克隆
组织表达谱
配体结合
荧光竞争结合实验
Spodoptera exigua
antennal binding protein
gene cloning
tissue expression pattern
ligand binding
fluorescence competitive binding assay
