白蛾周氏啮小蜂CcOBP5基因克隆、原核表达及其结合特性分析

Gene cloning,prokaryotic expression and analysis of binding characteristics of CcOBP5 of Chouioia cunea Yang

为了探究白蛾周氏啮小蜂气味结合蛋白CcOBP5与其寄主挥发物的结合特性,根据转录组测序数据,获得CcOBP5的全长CDNA序列,采用分子对接方法初步筛选能够与CcOBP5特异性结合的小分子化合物,通过原核表达和纯化CcOBP5蛋白,利用荧光结合实验检测重组CcOBP5与寄主挥发物的结合情况.结果显示,CcOBP5具有6个α螺旋和6个保守的半胱氨酸位点,能够与多种挥发性气味分子特异性结合,与美国白蛾蛹挥发物γ-丁内酯的结合最好,且γ-丁内酯对白蛾周氏啮小蜂有明显引诱作用.说明CcOBP5可能在白蛾周氏啮小峰对γ-丁内酯的行为反应中起关键作用.

To analyze the binding characteristics of the odorant binding protein 5(CcOBP5)in Chouioia cunea Yang with volatile compounds from the host,full-length cDNA sequence of CcOBP5 was obtained based on transcriptome sequencing data,after which molecular docking was employed to preliminarily screen small molecular compounds specifically binding to this protein.The recombinant CcOBP5 protein was expressed and purified in Escherichia coli,and then the protein was subjected to competitive fluorescence binding assay for verifying its combination with these volatile compounds.It was found that CcOBP5 has sixα-helix and six conservative L-Cysteine locations.CsOBP5 can specifically bind to a variety of volatile odor molecules,and there was a strong binding affinity between CcOBP5 andγ-Butyrolactone,which was a pupa volatile of Hyphantria cunea Drury.These results suggest thatCcOBP5may play a predominant role in the behavioral response of C.cunea toγ-Butyrolactone.