京水菜种子SU1抗菌蛋白的纯化、稳定性和抑菌机制  

Purification, stability and antifungal mechanism of antifungal protein SU1 from Brassica juncea var. multisecta seeds

作　　者：袁素素 秦武洒 叶秀娟 Yuan Susu;Qin Wusa;Ye Xiujuan(College of Plant Protection,Fujian Agricultural and Forestry University,Fuzhou 350002,Fujian Province,China)

机构地区：[1]福建农林大学植物保护学院,福州350002

出　　处：《植物保护学报》2019年第2期369-376,共8页Journal of Plant Protection

基　　金：福建省农业高校产学合作项目(2018N5005)

摘　　要：为拓展抗菌蛋白在新型生物农药上的潜在应用价值,通过SP-Sepharose阳离子交换层析、Mone S阳离子交换层析和Superdex 75凝胶过滤层析从京水菜Brassica juncea var. multisecta种子中纯化得到一种分子量约为30 kD的SU1抗菌蛋白,采用纸片扩散法测定该抗菌蛋白的稳定性和抑菌活性,并采用荧光染色法分析其抗菌机制。结果表明,在pH为1～3和11～13条件下,SU1抗菌蛋白抑菌活性不受影响;40～80℃热处理和100 mmol/L Mn^(2+)、Al^(3+)、Zn^(2+)、Cu^(2+)、Fe3+、Pb^(2+)、Mg^(2+)、K^+和Cr^(3+)离子处理后,该抗菌蛋白仍具有抑菌活性,而100 mmol/L Ca^(2+)离子处理后该抑菌蛋白的抑菌活性消失。此外,乙二胺四乙酸(ethylene diamine tetraacetic acid,EDTA)溶液处理后SU1抗菌蛋白的抑菌活性消失,再次加入Cr^(3+)离子时,其抑菌活性恢复。SU1抗菌蛋白能抑制7种植物病原真菌的生长,可破坏尖孢镰刀菌Fusarium oxysporum菌丝细胞膜透性和细胞核的完整性,并引起线粒体膜电势增加和菌丝细胞脱氧核糖核酸的分解。In order to increase the potential application value of antifungal proteins as new bio-pesticides, a 30-kD antifungal protein SU1 was purified from Brassica juncea var. multisecta seeds with cation exchange chromatography on SP-Sepharose, cation exchange chromatography on Mono S and gel filtration chromatography on Superdex 75 in this study. The stability and antifungal activity were measured by using filter paper method and antifungal mechanism of the protein SU1 was studied by using fluorescent staining method. The results showed that the antifungal activity of protein SU1 was fully preserved in the pH range of 1-3 and 11-13, temperature range of 40-80℃, and 100 mmol/L Mn2+,Al3+, Zn2+, Cu2+, Fe3+, Pb2+, Mg2+, K+ and Cr3+ ions. In contrast, the antifungal activity of protein SU1 disappeared following exposure to 100 mmol/L Ca2+ions. In addition, its antifungal activity disappeared after treatment with ethylene diamine tetraacetic acid(EDTA) solution, and its antifungal activity was reinstituted following exposure to Cr3+ ions again. The antifungal protein SU1 inhibited the growth of seven species of phytopathogenic fungi, permeabilized fungal membrane and disrupted nuclear integrity,disrupted fungal mitochondrial transmembrane potential and upregulated deoxyribonucleic acid(DNA)degradation in Fusarium oxysporum hyphae.

关 键 词：京水菜种子 抗菌蛋白 纯化 稳定性 抑菌机制 

分 类 号：TQ455[化学工程—农药化工]