嗜线虫致病杆菌HB310菌株杀虫蛋白的纯化及活性鉴定

摘要/Abstract

摘要：

嗜线虫致病杆菌Xenorhabdus nematophila HB310是从河北省土壤中筛选出的一株昆虫病原线虫体内分离纯化获得的共生菌，该菌的发酵液对多种昆虫有较高的杀虫活性。利用85％饱和度的硫酸铵盐析分别获得胞内蛋白提取物和上清液中胞外蛋白提取物，生测结果表明这两种蛋白提取物中都含有胃毒素和血腔毒素。通过制备型非变性凝胶电泳对蛋白提取物进行分离和纯化，得到了3种有杀虫活性的毒素蛋白（毒素Ⅰ、毒素Ⅱ和毒素Ⅲ），胞内的毒素蛋白与分泌到胞外上清液中的毒素蛋白是同种蛋白。毒素Ⅰ和毒素Ⅱ对棉铃虫初孵幼虫有明显的胃毒活性，但没有血腔毒性；毒素Ⅲ对大蜡螟幼虫有很强的血腔毒性，LD₅₀为0.18 μg/头。SDS-PAGE图谱显示毒素Ⅰ和毒素Ⅱ是由多个多肽组成的复合蛋白，而毒素Ⅲ只分离出一条多肽。毒素Ⅱ在50℃处理10 min，其杀虫活性没有显著变化；70℃处理10 min对毒素Ⅲ杀虫活性没有显著影响。

关键词: 嗜线虫致病杆菌, 毒素蛋白, 纯化, 杀虫活性

Abstract:

The extracts of extracellular and intracellular insecticidal proteins from Xenorhabdus nematophila HB310 strain, an entomopathogenic bacterium symbiotically associated with entomopathogenic nematode Steinernema carpocapsae, appeared high toxic activity by oral or injectable infection against several species of insects. Two oral insecticidal proteins, toxinⅠand toxinⅡ, and another haemocoel insecticidal protein, toxin Ⅲ with injectable activity were purified from the intracellular protein extract by native-PAGE. The same three protein toxins also were obtained from the extracellular protein extract. There was only a single band for any of toxins above, but the results of SDS-PAGE showed four bands for toxinⅠ, three bands for toxinⅡ and one band for toxin Ⅲ, respectively. Bioassay results indicated that the oral insecticidal activity of toxinⅡ was higher than that of toxinⅠ to Helicoverpa armigera neonates. The injectable hemocoelic potency (LD₅₀) of toxinⅢ was 0.18 μg/larva against Galleria mellonella fifth-instar larvae. The toxicity of toxinⅡ was stable under 50℃ in 10 min. The toxin Ⅲ was heat-stable to 70℃ in 10 min.

Key words: Xenorhabdus nematophila, protein toxins, purification, insecticidal activity

王勤英¹，南宫自艳¹，陆秀君¹，李秀花²，李国勋¹, 崔龙¹. 嗜线虫致病杆菌HB310菌株杀虫蛋白的纯化及活性鉴定[J]. , 2005, 48(3): 353-358.

WANG Qin-Ying¹, NANGONG Zi-Yan¹, LU Xiu-Jun¹, LI Xiu-Hua², LI Guo-Xun¹, CUI Long¹. Purification of insecticidal proteins from Xenorhabdus nematophila HB310 and detection of their insecticidal activity[J]. , 2005, 48(3): 353-358.

[1]	芦小鹏, 吴琼, 李毛龙, 李三兴, 马志卿, 张兴. 126种植物精油或精油主要成分对玉米黄呆蓟马的熏杀活性筛选[J]. , 2018, 61(8): 941-949.
[2]	陈秀琳, 苏丽, 陈丽慧, 李怡萍, 仵均祥, 李广伟. 梨小食心虫Minus-C气味结合蛋白的分子克隆、表达谱及结合特性分析[J]. , 2018, 61(7): 771-783.
[3]	张敏, 刘佳, 傅伟杰, 代光辉. 姜黄根提取物对苜蓿蚜成蚜有杀虫活性化合物的分离与鉴定[J]. , 2018, 61(6): 698-703.
[4]	张伟妮, 胡崇伟, 张梦馨, 韩威超, 黄小红. 意大利蜜蜂N-乙酰-β-D-氨基葡萄糖苷酶的分离纯化及性质分析(英文)[J]. , 2018, 61(10): 1153-1159.
[5]	赵静茹, 袁守丽, 高斌, 朱顺义. 抑制剂胱氨酸结多肽的抗菌和杀虫活性及进化[J]. , 2018, 61(10): 1212-1221.
[6]	苗娅, 张甜甜, 许柏英, 陈斌. 中华按蚊谷胱甘肽-S-转移酶E6（AsGSTE6）的表达纯化与结晶[J]. , 2018, 61(1): 59-67.
[7]	赵钰, 刘忠渊. 光滑鳖甲丝氨酸蛋白酶抑制剂基因ApSerpin-FA72的克隆、表达及功能验证[J]. , 2017, 60(8): 857-864.
[8]	谭永安, 赵旭东, 郝德君, 肖留斌, 柏立新, 赵静, 孙洋, 姜义平. 绿盲蝽磷脂酶C(AlPLC)的表达、纯化及酶学性质[J]. , 2017, 60(11): 1247-1254.
[9]	房迟琴, 张鑫鑫, 刘丹丹, 李克斌, 张帅, 曹雅忠, 樊东, 尹姣. 暗黑鳃金龟气味结合蛋白HparOBP15a基因的克隆及功能分析[J]. 昆虫学报, 2016, 59(3): 260-268.
[10]	海力且木·艾力, 徐鑫, 刘忠渊. 光滑鳖甲抗菌肽的原核表达条件优化及其抗菌活性[J]. 昆虫学报, 2016, 59(1): 8-15.
[11]	Tolulope Felix ADEDIRAN, Isaac Olusanjo ADEWALE. 臭腹腺蝗肠道中共存的淀粉和纤维素消化酶的纯化、动力学和抑制研究（英文）[J]. , 2015, 58(7): 739-749.
[12]	李尚伟, 赵柏松, 杜娟. 九香虫抗菌肽CcAMP1的分离纯化和抗菌活性检测[J]. , 2015, 58(6): 610-616.
[13]	陈伟, 徐卫华. 棉铃虫Wnt-1基因克隆、多克隆抗体制备及在滞育和非滞育蛹脑中的表达检测[J]. , 2015, 58(1): 15-21.
[14]	赵同伟，张兵，吕文静，翟会锋，吴桃燕，张石洋，汤小伟，冯从经. 亚洲玉米螟酚氧化酶原的原核表达和性质分析[J]. , 2014, 57(7): 798-805.
[15]	孙海霞，陈俊，杨之帆. 褐飞虱细胞色素P450基因CYP4C62的原核表达及多克隆抗体的制备[J]. , 2014, 57(6): 656-662.

嗜线虫致病杆菌HB310菌株杀虫蛋白的纯化及活性鉴定

Purification of insecticidal proteins from Xenorhabdus nematophila HB310 and detection of their insecticidal activity

PDF (PC)

赞

可视化

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 15

Metrics

本文评价

推荐阅读 10