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

Abstract: 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-tetradecenol (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.

Key words: Spodoptera exigua, antennal binding protein, gene cloning, tissue expression pattern, ligand binding, fluorescence competitive binding assay