绛红褐链霉菌YSSPG3的鉴定及其抗菌蛋白纯化  

Identification of Streptomyces purpeofuscus YSSPG3 and purification of its antifungal protein

作　　者：张丽娜[1] 朱天辉[2] 李芳莲[2] 

机构地区：[1]四川农业大学农学院,雅安625014 [2]四川农业大学林学院,雅安625014

出　　处：《植物病理学报》2012年第3期242-251,共10页Acta Phytopathologica Sinica

基　　金：国家自然科技资源共享平台(2005DKA21207-13);长江上游林业生态工程省级重点实验室资助项目

摘　　要：从四川碧峰峡撑×绿杂交竹健康植株根际分离到对杂交竹梢枯病菌(Arthrinium phaeospermum)有强拮抗作用的放线菌菌株YSSPG3,该菌株的无菌发酵液对多种真菌和细菌具有抗菌活性。通过对菌株YSSPG3的形态特征、培养特性、生理生化特性、细胞壁组分与16S rDNA序列分析,确定为绛红褐链霉菌(Streptomyces purpeofuscus)。采用硫酸铵沉淀和柱层析法对发酵液中起主要抑菌作用的抗菌物质进行分离纯化,获得单一的抗菌蛋白AMP,分子量为5 075.619 Da,等电点为6.77。抗菌蛋白在温度≥60℃和pH<7,以及紫外线照射时间≥12 h的环境下抑菌活性均明显下降,但受蛋白酶K、胰蛋白酶和氯仿影响不大,无几丁质酶、葡聚糖酶等水解酶活性。利用Edman降解法测定抗菌蛋白N末端25个氨基酸序列,与来自Streptomyces tendae Tü901的chitin-binding protein AFP1的同源性较高,为81.00%。The actinomyces strain YSSPG3 suppressing Arthrinium phaeospermum was isolated from rhizosphere of healthy hybrid bamboo in Sichuan,and its fermentation filtrate showed inhibitory effect to many fungi and bacteria.Based on the study of its morphological,physiological and biochemical characteristics,together with sequencing of 16S rDNA genes and phylogenetic analysis,strain YSSPG3 was identified as Streptomyces purpeofuscus.The antifungal protein(AMP) with a molecular mass of 5 075.619 Da was purified by the methods of ammonium sulfate precipitating and column chromatography.The isoelectric point(pI) of AMP was 6.77 and lacked the activity of hydrolytic enzyme such as chitinase and glucanase.The inhibitory activity of the antifungal protein decreased distinctly at the higher temperature(≥60℃),in the condition of acidity(pH<7) and a long ultraviolet radiation time(≥12 h),but was tolerant of proteinase K,trypsin and chloroform.The N-terminal sequence between the antifungal protein and a kind of chitin-binding protein AFP1 from S.tendae Tü901 was of higher homology in 81.00%.

关 键 词：绛红褐链霉菌 杂交竹梢枯病菌 抗菌蛋白 分离纯化 N端序列 

分 类 号：S432.23[农业科学—植物病理学]