Xu Xiaoli, Huang Jinming, Fang Jun, Lin Chaoyang, Cheng Jiaan, Shen Zhicheng
Institute of Insect Sciences and National Key Laboratory of Rice Biology, School of Agriculture and Biotechnology, HuajiaChi Campus, Zhejiang University, Hangzhou 310029, China.
Protein Expr Purif. 2008 Oct;61(2):113-6. doi: 10.1016/j.pep.2008.05.019. Epub 2008 Jun 8.
Glucoamylase, which catalyses the hydrolysis of the alpha-1,4 glycosidic bonds of starch, is an important industrial enzyme used in starch enzymatic saccharification. In this study, a glucoamylase gene from Aspergillus awamori, under the control of the promoter of seed storage protein Gt1, was introduced into rice by Agrobacterium-mediated transformation. Significant glucoamylase activity was detected specifically in the seeds but not other tissues of the transgenic rice lines. The highest enzymatic activity was found in the transgenic line Bg17-2, which was estimated to have about 500 units per gram of seeds (one unit is defined as the amount of enzyme that produces 1 micromol of reducing sugar in 1 min at 60 degrees C using soluble starch as substrate). The optimum pH for the activity of the rice produced enzyme is 5.0-5.5, and the optimum temperature is around 60 degrees C. One part of this transgenic glucoamylase rice seed flour fully converted 25 parts of corn starch pre-liquefied by an alpha-amylase also produced by a transgenic rice into glucose in 16 h incubation. This study suggests that this hydrolysis enzyme may substitute commercial fermentation enzymes for industrial starch conversion.
糖化酶可催化淀粉的α-1,4糖苷键水解，是淀粉酶解糖化过程中一种重要的工业用酶。在本研究中，来自泡盛曲霉的糖化酶基因在种子贮藏蛋白Gt1启动子的控制下，通过农杆菌介导转化法导入水稻。在转基因水稻株系的种子中特异性检测到显著的糖化酶活性，而在其他组织中未检测到。在转基因株系Bg17-2中发现了最高的酶活性，估计每克种子约有500个酶活力单位（一个酶活力单位定义为在60℃下以可溶性淀粉为底物，每分钟产生1微摩尔还原糖的酶量）。水稻产生的该酶活性的最适pH为5.0 - 5.5，最适温度约为60℃。将一份这种转基因糖化酶水稻种子粉与25份同样由转基因水稻产生的α-淀粉酶预液化的玉米淀粉混合，在16小时的孵育后可完全转化为葡萄糖。本研究表明，这种水解酶可能替代商业发酵酶用于工业淀粉转化。
