植物抗逆蛋白(LEA3)22-氨基酸耐盐结构域在酵母细胞中的鉴定  

Functional Identification of Plant Stress-resistant Protein(LEA3) and Its 22-mer Motifs in Yeast

作　　者：刘昀[1] 李冉辉[1] 汪为茂[1] 梁翔宇[1] 郑易之[1] 

机构地区：[1]深圳大学生命科学学院,深圳市微生物基因工程重点实验室,深圳518060

出　　处：《植物研究》2009年第1期74-79,共6页Bulletin of Botanical Research

基　　金：国家自然科学基金(30670180和30470107)

摘　　要：大豆PM2蛋白属LEA3(late embryogenesis abundant3)蛋白。本文构建了编码PM2全长及含22-氨基酸结构域多肽(PM2、PM2A、PM2B和PM2C)的酵母表达载体。转化酵母得到四种重组菌。SDS-PAGE电泳和ESI-MS/MS或MALDI-TOF/TOF质谱结果表明,重组菌可表达目标多肽。测定对照菌及重组菌在无胁迫、高盐(1.5mol.L-1NaCl)和高渗透(2mol.L-1山梨糖)下的生长曲线。结果表明,在高盐胁迫下,四种重组菌胁迫后的恢复明显好于对照菌。多肽对高盐耐受能力的大小为:PM2C>PM2B≈PM2A≈PM2。证明大豆PM2蛋白的表达可提高酵母耐盐性,且22-氨基酸基序为PM2蛋白的耐盐结构域。结合前文在大肠杆菌中的结果,为"LEA蛋白可以类似机制参与原核和真核生物耐盐保护作用"的假说提供实验支持。然而,在高山梨糖胁迫下,对照菌和酵母重组菌的生长情况无明显差异。Soybean PM2 protein belongs to the family of group 3 LEA （late embryogenesis abundant） proteins. In the present paper, yeast expression vectors encoding full-length PM2 protein and deleted polypeptides containing 22-mer motif were constructed. The yeast recombinants were obtained after transformation. Expression of PM2 and the deleted polypeptides of PM2A, PM2B and PM2C were identified in yeast lysis by SDS-PAGE and ESI-MS/MS or MALAI-TOF/TOF analysis. The growth performances of yeast recombinants and their control with empty vector were tested under normal, high salinity（ 1.5 mol · L^-1 NaCl）or high osmotic（2 mol · L^-1 sorbitol） conditions. The results showed that ： 1 ） the expressions of PM2 protein and its 22-mer motif are not deleterious to the growth behavior of yeast recombinants under normal condition ; 2） the recoveries of 4 recombinants are better than that of control with the empty vector under high salinity conditions, indicating that PM2 protein and its 22- mer motif could improve salt tolerance of yeast directly. The high salt tolerance of yeast conferred by 4 polypeptides are PM2C 〉 PM2B ≈ PM2A ≈ PM2; 3 ） no growth difference was observed between the 4 reeombinants and the control under high sorbitol stress. These results showed that 22-met motif is a functional domain in PM2 protein and it is consist with the results described as in E. coli. All these results provide directly experimental support for the hypothesis that ＂LEA proteins may represent analogous protection to the high salinity stress in prokaryotes and eukaryotes＂ for the first time.

关 键 词：LEA3蛋白 PM2蛋白 耐盐性 酵母 22-氨基酸基序 

分 类 号：S565.1[农业科学—作物学]