中红侧沟茧蜂气味结合蛋白MmedOBP18的原核表达及配体结合特性  

Prokaryotic expression and ligand binding characteristics of odorant binding protein MmedOBP18 of the parasitoid wasp Microplitis mediator (Hymenoptera: Braconidae)

作　　者：宋玄 单双 王山宁 陶宇逍 李瑞军[1] 张永军[2] SONG Xuan;SHAN Shuang;WANG Shan-Ning;TAO Yu-Xiao;LI Rui-Jun;ZHANG Yong-Jun(College of Plant Protection,Agricultural University of Hebei,Baoding,Hebei 071000,China;State Key Laboratory for Biology of Plant Diseases and Insect Pests,Institute of Plant Protection,Chinese Academy of Agricultural Sciences,Beijing 100193,China;College of Plant Protection,China Agricultural University,Beijing 100193,China;Institute of Plant and Environment Protection,Beijing Academy of Agriculture and Forestry Sciences,Beijing 100097,China)

机构地区：[1]河北农业大学植物保护学院,河北保定071000 [2]中国农业科学院植物保护研究所植物病虫害生物学国家重点实验室,北京100193 [3]中国农业大学植物保护学院,北京100193 [4]北京农林科学院植保护环境保护研究所,北京100097

出　　处：《昆虫学报》2019年第2期150-159,共10页Acta Entomologica Sinica

基　　金：国家自然科学基金项目(31672038;31621064)

摘　　要：【目的】研究中红侧沟茧蜂Microplitis mediator气味结合蛋白MmedOBP18在触角中的表达定位,解析MmedOBP18重组蛋白的配体结合特性。【方法】原核表达中红侧沟茧蜂气味结合蛋白MmedOBP18;采用荧光免疫组织化学技术研究MmedOBP18在中红侧沟茧蜂雌蜂触角中的表达定位;通过荧光竞争结合实验分析MmedOBP18重组蛋白与99种候选配体的结合特性。【结果】在原核表达系统中成功表达MmedOBP18重组蛋白。荧光免疫定位结果显示MmedOBP18主要表达在触角锥形感器Ⅰ内的淋巴液中。荧光竞争结合实验结果表明,MmedOBP18重组蛋白能够与16种候选配体结合,与低挥发性植物挥发物2-十三酮、十二醛、十四酸和十一酸的结合能力最强,其解离常数K_i值分别为5.21, 6.42, 6.49和6.58μmol/L;而且MmedOBP18重组蛋白与非挥发性植物次生物质棕榈酸、棉酚、槲皮素及亚麻酸也有较强的结合能力,K_i值分别为3.86, 5.07, 5.08和6.51μmol/L;此外,MmedOBP18重组蛋白与鳞翅目昆虫性信息素组分顺-9-十四碳烯醛及顺-11-十六碳醛表现出较强的结合能力,K_i值分别为9.09和11.67μmol/L,提示该蛋白在寄主定位过程中可发挥重要的作用。【结论】中红侧沟茧蜂气味结合蛋白MmedOBP18能够结合长链低挥发和非挥发性化合物,推测其在嗅觉和味觉识别中发挥双重功能,主要参与对寄主和寄主生境的化学信息的近距离识别。【Aim】To clarify the expression of the odorant binding protein MmedOBP18 in the antenna of Microplitis mediator,and to investigate the ligand binding characteristics of the recombinant MmedOBP18.【Methods】The recombinant MmedOBP18 of M.mediator was expressed in the prokaryotic expression system.The localization of expression of MmedOBP18 in the antenna of female wasps of M.mediator was evaluated using fluorescence immunohistochemistry assay.The binding characteristics of the recombinant MmedOBP18 protein to 99 candidate ligands were analyzed by fluorescence competitive binding assay.【Results】The recombinant MmedOBP18 protein was successfully expressed in the prokaryotic expression system.The immunofluorescence localization result showed that MmedOBP18 was mainly expressed in the lymph of sensilla basiconica type I of the antenna.The recombinant MmedOBP18 could strongly bind with 16 kinds of ligands.Among the low volatile compounds,2-tridecanone,dodecyl aldehyde,myristic acid and undecylic acid showed the strongest binding capacities to the recombinant MmedOBP18,with the dissociation constant(K i)values of 5.21,6.42,6.49 and 6.58μmol/L,respectively.The non-volatile compounds palmitic acid,gossypol,quercetin and linolenic acid also showed strong binding capabilities to the recombinant MmedOBP18,with the K i values of 3.86,5.07,5.08 and 6.51μmol/L,respectively.In addition,the lepidopteran pheromone components(Z)-9-tetradecenal and(Z)-11-hexadecenal also showed strong binding capabilities to MmedOBP18,with the K i values of 9.09 and 11.67μmol/L,respectively,suggesting an important role of MmedOBP18 in host localization.【Conclusion】The MmedOBP18 of M.mediator has strong binding capabilities to long-chain low volatile or non-volatile compounds.We speculated that MmedOBP18 may play a dual role in olfactory and taste recognition,and is mainly involved in the close range perception of chemical signals from host and host habitats.

关 键 词：中红侧沟茧蜂 气味结合蛋白 MmedOBP18 荧光免疫定位 配体结合特性 化学感受 

分 类 号：Q966[生物学—昆虫学]