亚洲玉米螟CPT1蛋白的表达、纯化和与几丁质的结合能力  

Expression,Purification and Characterization of Chitin-binding in Protein CPT1 from Ostrinia furnacalis

作　　者：蒋望 包涵 陈金利 刘田[1] 杨青[1,2,3] JIANG Wang;BAO Han;CHEN Jinli;LIU Tian;YANG Qing(School of Bioengineering,Dalian University of Technology,Dalian 116024,China;Institute of Plant Protection,Chinese Academy of Agricultural Sciences,Beijing 100193,China;Agricultural Genomics Institute at Shenzhen,Chinese Academy of Agricultural Sciences,Shenzhen 518000,China)

机构地区：[1]大连理工大学生物工程学院,大连116024 [2]中国农业科学院植物保护研究所,北京100193 [3]中国农业科学院农业基因组研究所,深圳518000

出　　处：《中国生物防治学报》2023年第2期462-470,共9页Chinese Journal of Biological Control

基　　金：国家重点研发计划(2017YFD0200500,2018YFD0200100);国家自然科学基金(31871959)。

摘　　要：亚洲玉米螟Ostrinia furnacalis是为害我国主要农作物玉米的一种重要害虫。昆虫表皮在运动、抵御机械损伤及对抗外界环境压力等方面发挥关键性作用。Tweedle(CPT)家族表皮蛋白在形成昆虫表皮中发挥重要作用,研究CPT有助于开发新一代害虫防治方法。本研究在大肠杆菌系统中以包涵体的形式重组表达了Of CPT1蛋白,使用金属螯合层析进行纯化,复性得到了Of CPT1蛋白,收率为50 mg/L。几丁质结合试验发现,Of CPT1对α-几丁质、β-几丁质、壳聚糖、胶体几丁质和几丁质纳米微晶均有一定的结合能力,其中与几丁质纳米微丝结合能力最强。圆二色光谱结果表明,Of CPT1蛋白与壳聚糖结合后二级结构从以α-螺旋占主导变为以β-折叠占主导。本研究利用原核表达系统获得了亚洲玉米螟重组CPT蛋白并表征了它的部分生物化学性质。Asian corn borer,Ostrinia furnacalis,is an important agricultural insect pest that greatly impairs corn production in China.Insect cuticle plays key roles in protecting insects from locomotion,mechanical damage or environmental stresses.The cuticle proteins that belong to Tweedle(CPT)family are known to play important roles in insect cuticle formation.Investigation on CPT may help the development of new methods for insect pest control.In this study,OfCPT1,one of the CPTs derived from the Asian corn borer,was recombinantly expressed as inclusion bodies in Escherichia coli.The obtained OfCPT1 was purified by metal chelate chromatography and then renatured with a yield of 50 mg/L.Chitin binding assay showed that OfCPT1 had a strong binding ability with chitin nanofibers,but moderate binding toα−chitin,β−chitin,chitosan,colloid chitin,and chitin nanocrystal.The circular dichroism spectra of OfCPT1 before and after binding with chitosan,showed a structural change fromα−helix dominance toβ−sheet dominance.This work has accomplished the recombinant preparation of OfCPT1 and revealed its biochemical properties for the first time.

关 键 词：表皮蛋白 Tweedle 几丁质结合 杀虫剂靶标 

分 类 号：S476[农业科学—农业昆虫与害虫防治]