芥子酶是广泛存在于十字花科植物中的一类降解硫代葡萄糖苷的酶。当植物受到病虫侵袭时，
芥子酶与其底物硫代葡萄糖苷结合，释放有毒的水解产物形成植物重要的化学防御系统。TGG5 是新
发现的一类芥子酶基因，对其研究将有利于揭示芥子酶防御系统的功能。试验克隆了拟南芥TGG5 基
因cDNA，构建酵母表达载体，转化毕氏酵母GS115。经甲醇诱导表达和Ni 亲和层析，获得纯化的
TGG5 重组蛋白。SDS-PAGE分析表明，重组蛋白分子量集中在58 kD左右，并有较明显拖尾，呈现典型
的糖基化蛋白的特征。进一步探讨了TGG5 重组蛋白的酶学特性，结果表明：该芥子酶的Km为1.96
mmol/L，Vmax为0.084 mmol/(min·mg)；酶反应的最适pH 为5.5；该芥子酶在37~80℃范围内有较广泛的
温度适应性，并具有较好的80℃热稳定性；NaCl对该酶有明显的抑制作用；低浓度抗坏血酸具有激活芥
子酶的作用，抗坏血酸终浓度为1.5 mmol/L时，酶活力最大。RT-RCR分析，证实了TGG5 只在拟南芥根
中特异表达，这表明TGG5 可能代表了一个芥子酶基因新亚家族。

Myrosinase widely exists in the Cruciferae plant, which can catalyse the hydrolysis of
thioglucosides. Myrosinase and glucosinolate are mixed upon infestation by pests, and thioglucosides are
hydrolyzed into toxic compounds to serve as defense weapons. A new myrosinase gene was found, denoted as
TGG5 . The research of myrosinase geneTGG5 could explore the function of myrosinase defense system. In this
research, the cDNA sequence of TGG5 was cloned, and the pichia expression vector pPIC3.5K was
constructed, then transformation of pichia GS115 was made. The expression of recombinantTGG5 protein was
induced by Methanol, and purified by Ni-chelating colum. SDS-PAGE analysis showed that the recombinant
protein concentrated in the 58 kD molecular weight around and showed typical features of protein glycosylation
with a more visible tail. The myrosinase activity of recombinantTGG5 protein was future researched after
separation and purification. The results indicated that theK m andV max of the enzyme were 1.96 mmol/L and
0.084 mmol/(min·mg)respectively. The optimal pH of tested myrosinase was 5.5. The myrosinase showed the
wider adaptability in the temperature range of 37~80℃, and took on better stability on 80℃ high temperature.
NaCl had an obvious inhibitory effect on activity of tested myrosinase, but low concentration of ascorbic acidcould activate the myrosinase, which reached to a climax with an ascorbic acid final concentration of 1.5 mmol/L.
RT-RCR detection confirmed the TGG5 gene only specificlly expressed in roots, which showed that TGG5
represent a novel myrosinase gene subfamily.