苏云金芽孢杆菌几丁质酶Chi73的特性及生物活性研究  

Characteristics and biological activity of the chitinase Chi73 from Bacillus thuringiensis

作　　者：刘佳[1] 宋萍[1] 张杰 南宫自艳[1] 王勤英[1] LIU Jia;SONG Ping;ZHANG Jie;NANGONG Ziyan;WANG Qinying(College of Plant Protection, Hebei Agricultural University, Baoding 071000, China;Luanping State-owned Forestry Farm Management of Chengde City, Chengde 068250, China)

机构地区：[1]河北农业大学植物保护学院,河北保定071000 [2]河北省承德市滦平国有林场管理处,河北滦平068250

出　　处：《河北农业大学学报》2019年第4期74-81,共8页Journal of Hebei Agricultural University

基　　金：十三五国家重点研发计划课题(2017YFD0300906)

摘　　要：为了明确苏云金芽孢杆菌(Bacillus thuringiensis, Bt)几丁质酶Chi73的特性和功能,本研究从Bt kurstaki HD-73菌株中克隆了几丁质酶基因chi73,并对其进行了原核表达;随后对表达的重组几丁质酶Chi73的酶学特性、杀虫活性和抑菌活性开展了研究。结果表明,在大肠杆菌中正确表达了大小为77 kDa的重组几丁质酶Chi73。以胶体几丁质为底物时,该酶的最适反应温度为60℃,最适pH为7.0,Zn2+和SDS对酶活力有明显的抑制作用。Chi73能显著抑制棉铃虫2龄幼虫的生长,其抑制率为94.37%。同时,对Bt Cry1Ac毒素有明显的增效作用,使Bt Cry1Ac毒素对棉铃虫2龄幼虫的LC50由27.39μg/mL降低至14.49μg/mL。此外,Chi73对白菜黑斑病菌、葡萄白腐病菌和棉花枯萎病菌均表现出明显的抑制作用。可见,Bt几丁质酶在杀虫增效和抑制真菌方面具有一定的应用潜力。In order to clarify the characteristics and function of the chitinase Chi73 from Bacillus thuringiensis kurstaki HD-73, the chi73 gene of Bt kurstaki HD73 was cloned and expressed in Escherichia coli. Then, the enzymatic activity, insecticidal activity and antifungal activity of recombinant chitinase Chi73 were studied. The results showed that the recombinant chitinase Chi73 was successfully expressed in E. coli BL21(DE3). The optimum temperature and pH of Chi73 was 60 ℃ and pH 7.0, respectively, whereas Zn2+ and SDS exhibited inhibition on the activity of Chi73. Moreover, Chi73 could significantly inhibit the growth of the second instar larva of Helicoverpa armigera with a growth inhibition rate of 94.37%. At the same time, the Chi73 had obvious synergistic effect with the insecticidal activity of Bt Cry 1Ac. After adding Chi73, the LC50 of Bt Cry1Ac toxin against the second instar larva of H. armigera was reduced from 27.39 μg/mL to 14.49 μg/mL. Besides, the Chi73 showed antifungal activity against Alternaria brassicicola, Coniothyrium diplodiella and Fusarium oxysporum. The present study provides that chitinase of Bt has the potential in insecticidal synergy effect and pathogenic fungi inhibition.

关 键 词：苏云金芽孢杆菌 几丁质酶 酶活性 杀虫活性 抑菌活性 

分 类 号：S476.1[农业科学—农业昆虫与害虫防治]