耐热古菌芝田硫化叶菌海藻糖生成相关酶的基因克隆、表达和序列分析  

Cloning, Expression and Sequence Analysis of a Cluster of Genes Encoding New Trehalose-producing Enzymes From Thermophilic Archaebacterium Sulfolobus shibatae B12

作　　者：吴襟[1] 于炜婷[1] 王辉[1] 刘莉[1] 王绍校[1] 张树政[1] 

机构地区：[1]中国科学院微生物研究所,北京100080

出　　处：《生物化学与生物物理进展》2003年第5期798-802,共5页Progress In Biochemistry and Biophysics

基　　金：北京市自然科学基金课题(5 982 0 10 );国家高技术 863 "计划资助项目(2 0 0 1AA2 14 14 1)~~

摘　　要：从耐热古菌海藻糖芝田硫化叶菌B12中分别克隆出海藻糖生成相关酶———麦芽寡糖基海藻糖合酶的基因treY和麦芽寡糖基海藻糖基水解酶的基因treZ,测定了其核苷酸序列并进行了表达.其中treY编码的蛋白质有 72 8个氨基酸、分子质量为 86ku;treZ编码的蛋白质有 5 5 9个氨基酸、分子质量为 6 5ku.它们与已报道的其他微生物的两个海藻糖生成相关酶的基因进行同源性比较,treY和treZ的同源性分别为 93%和 76 % (硫矿硫化叶菌P2)、97%和 95 % (硫矿硫化叶菌KM1)、6 3%和 6 6 % (嗜酸热硫化叶菌ATCC3390 9)、4 8%和 5 0 % (节杆菌Q36 )、4 8%和 5 2 %(根瘤菌M11)、5 0 %和 5 2 %(短杆菌).通过PHYLIP软件进行这些基因序列的分类聚类计算,获得这几种微生物间两个酶类的蛋白质系统进化树;经过氨基酸序列比较分析还发现,所有的海藻糖生成相关酶都含有糖苷酶家族 13中几个高度保守的α 淀粉酶催化活性区。Two genes (treY and treZ) encoding for trehalose-producing enzymes, a maltooligosyl trehalose synthase (MTSase) and a maltooligosyl trehalose trehalohydrolase (MTHase), had been cloned, expressed and sequenced from thermophilic archaebacterium Sulfolobus shibatae B12. The nucleotide sequences of treY and treZ indicated proteins with lengths of 728 and 559 amino acids and molecular masses of 86 ku and 65 ku, respectively. treY and treZ genes from S. shibatae B12 show high sequence homologies with other microbial MTSase and MTHase genes, such as S. solfataricus P2 (93% and 76% identity), S. solfataricus KM1 (97% and 95% identity), Sulfolobus acidocaldarius ATCC 33909 (63% and 66% identity), Arthrobacter sp. Q36 (48% and 50% identity), Rhizoblum sp. M-11 (48% and 52% identity), Brevibacterium sp. (50% and 52% identity). Phylogenetic trees of these enzymes genes were constructed by analysis class of their overall nucleotide sequences. The amino acid sequences of MTSase and MTHase showed identity with members of the glycosyl hydrolase family 13 (alpha-amylase family). Four important regions highly conserved in the glycosyl hydrolase family 13 exist in the amino acid sequences of all these trehalose-producing enzymes. It is suggested that MTSase and MTHase are members of glycosyl hydrolase family 13 and treY and treZ maybe derived from a common original alpha-amylase gene.

关 键 词：耐热古菌 芝田硫化叶菌 海藻糖 酶 基因克隆 表达 序列分析 

分 类 号：Q936[生物学—微生物学] Q78