类芽孢杆菌Bg1抗真菌蛋白的分离及其特性分析  

Purification and Characteristics Analysis of the Antifungal Protein Produced by Paenibacillus sp. Bg1

作　　者：叶慧敏[1,2] 张殿朋[2] 刘霞[1] 于莉[1] 刘伟成[2] 

机构地区：[1]广东海洋大学农学院,广东湛江524088 [2]北京市农林科学院植物保护环境保护研究所,北京100097

出　　处：《核农学报》2014年第1期29-35,共7页Journal of Nuclear Agricultural Sciences

基　　金：广东省科技厅项目(2009B060600008);北京市科技计划课题(Z121100001212002);北京市农林科学院科技创新基金(KJCX201101001)

摘　　要：为了明确类芽孢杆菌（Paenibacillus sp．）Bg1的主要抗真菌物质及其特性，本文利用抑菌圈法测定了Bg1抗菌蛋白在各种处理条件下抑菌活性的稳定性；通过60％硫酸铵沉淀、DEAE—Sephadex A50离子交换层析及Sephadex G100凝胶过滤层析进行了抗菌蛋白的分离纯化，以SDS-PAGE进行了其纯度检测。结果表明，28～100℃处理30min、4℃存放55d及在pH值3～7范围内菌株Bg1抗菌蛋白抑菌活性均无显著性变化，但pH值〉8显著影响其抑菌活性，蛋白酶处理则使其抑菌活性完全丧失；纯化后所得抗菌蛋白分子量约10KD。据此可知，类芽孢杆菌Bg1主要抗菌活性物质为低分子量蛋白，其对热和酸性条件及贮存时间稳定性好，具有进一步开发应用的潜能。In order to identify the key antifungal substance of Paenibacillus sp. Bgl and its main characteristics, the separation, purification and the stability analysis of the antifungal protein produced by strain Bgl was carried out. To determine the active stability of the antifungal protein under various processing conditions, a method by measuring the inhibition zone was applied. The cell culture of Pctenibacillus sp. Bgl was obtained by shake-flask fermenting. The crude proteins were extracted by precipitating with 60% saturation （NH4 ）2SO4. Then the crude protein was separated and purified by DEAE-Sephadex A50 ion-exchange column chromatography and Sephadex G100 gel filtration column chromatography. The purified active protein was detected by SDS-PAGE. The results indicated that the crude protein of strain Bgl showed no significant changes in the antifungal activity under the following treatment conditions： 28℃ -100℃for 30 min, 4℃ for 55 d and pH 3 - 7. But the antifungal activity was significantly affected by the alkaline conditions of pH 〉8 and completely lost by treatment with protease. After three-step purification, the purified active protein was demonstrated to be a single protein band with the molecular weight of 10 KD by SDS-PAGE. It was concluded that the main antifungal substance of Paenibacillus sp. Bgl belongs to the low molecular weight protein which presented a good stability towards heat, acidity and storage time, and exhibited a huge potential for further development and application.

关 键 词：类芽孢杆菌 抗真菌蛋白 分离纯化 稳定性分析 

分 类 号：S432.4[农业科学—植物病理学]