关键词: 桔小实蝇, 气味结合蛋白, 组织表达谱, 原核表达, 荧光竞争结合实验

Abstract: In order to study the function and binding characteristics of the odorant-binding proteins (OBPs) in the oriental fruit fly, Bactrocera dorsalis, we cloned an OBP cDNA from B. dorsalis, which was named BdorOBP2 (GenBank accession no.: KC773766), and then heterologously expressed its protein in prokaryotic cells. BdorOBP2 contains a 447-bp open reading frame (ORF) encoding 148-amino-acid residues. The mature protein of BdorOBP2 includes six conserved cysteine residues, which are the hallmark of insect OBPs. Real-time quantitative PCR results showed that BdorOBP2 was expressed in different tissues with the highest expression in the head and the lowest in the wings (63%±6% of the expression level in the head). The recombinant protein BdorOBP2 was purified by affinity chromatography, and N-phenyl-1-naphthylamine (1-NPN) was applied as a fluorescent probe to measure the binding capacity of BdorOBP2 with each of seven major host fruit odors. The results indicated that BdorOBP2 can evidently bind to most esters and aldehydes examined. BdorOBP2 showed the highest affinity to trans-2-hexenal and β-ionone with the dissociation constant KD of 9.96 and 15.37 μmol/L, respectively. The results of this study may help rational designs of specific and effective attractants to be used for managing B. dorsalis.

Key words: Bactrocera dorsalis, odorant-binding protein (OBP), tissue expression pattern, prokaryotic expression, fluorescence competitive binding assay