芝田硫化叶菌麦芽寡糖基海藻糖合酶基因在大肠杆菌中的克隆和表达

用ＰＣＲ 方法从芝田硫化叶菌中扩增了编码一种新酶，即麦芽寡糖基海藻糖合酶（ ＭＴＳａｓｅ） 的基因，扩增的２－２ｋｂ ＤＮＡ 插入到原核表达载体ｐＢＶ２２０ 中，构建成重组质粒ｐＳＢＧＴ１ 。ｐＳＢＧＴ１ 中ＭＴＳａｓｅ 基因在大肠杆菌中得到表达。ＳＤＳＰＡＧＥ 分析表达产物ＭＴＳａｓｅ蛋白的分子量约为７４ｋＤａ ，同核苷酸序列测定所推导的值相符。表达产物占细胞总蛋白约４－４ ％ 。ｐＳＢＧＴ１ 产生的重组酶作用于淀粉部分水解物，使ＤＥ 值降低，得到非还原糖或低还原糖。

CLONING AND EXPRESSION OF THE GENE ENCODING MALTOOLOGOSYL TREHALOSE SYNTHASE FROM SULFOLOBUS SHIBATAE IN E. COLI

2.2 kb DNA fragment encoding a novel enzyme, maltooligosyl trehalose synthase (MTSase)was amplified from Sulfolobus shibatae by using PCR technique. The amplified 2.2kb DNA fragment was inserted into an expression vector, pBV220, to yield the recombinant plasmid pSBGT1. MTSase gene in pBSGT1 was expressed in E. coli. The molecular weight of expressed MTSase detected by SDS-PAGE was about 74kD, which is conformed with that deduced from nucleotide sequence. The expressed MTSase protein accounted for about 4.4% of the total cell protein. The MTSase from transformants containing pBSGT1 is capable of decreasing DE value, forming non-reducing or less-reducing saccharides when allowed to act on reducing partial starch hydrolysates.