Tokunaga M, Ishibashi M, Tatsuda D, Tokunaga H
Laboratory of Applied Microbiology, Faculty of Agriculture, Kagoshima University, Japan.
Yeast. 1997 Jun 30;13(8):699-706. doi: 10.1002/(SICI)1097-0061(19970630)13:8<699::AID-YEA124>3.0.CO;2-N.
We constructed two mouse alpha-amylase secretion vectors for Kluyveromyces lactis using the well-characterized signal sequence of the pGKL 128 kDa killer precursor protein. Both PHO5 and PGK expression cassettes from Saccharomyces cerevisiae directed the expression of mouse alpha-amylase in YPD medium at a similar level of efficiency. K. lactis transformants secreted glycosylated and non-glycosylated alpha-amylase into the culture medium and both species were enzymatically active. The K. lactis/S. cerevisiae shuttle secretion vector pMI6 was constructed, and K. lactis MD2/1(pMI6) secreted about four-fold more alpha-amylase than S. cerevisiae YNN27 harboring the same plasmid, indicating that K. lactis is an efficient host cell for the secretion and production of recombinant proteins.
我们使用特征明确的pGKL 128 kDa杀伤前体蛋白信号序列构建了两个用于乳酸克鲁维酵母的小鼠α-淀粉酶分泌载体。来自酿酒酵母的PHO5和PGK表达盒在YPD培养基中指导小鼠α-淀粉酶的表达，效率水平相似。乳酸克鲁维酵母转化体将糖基化和非糖基化的α-淀粉酶分泌到培养基中，且两种形式均具有酶活性。构建了乳酸克鲁维酵母/酿酒酵母穿梭分泌载体pMI6，与携带相同质粒的酿酒酵母YNN27相比，乳酸克鲁维酵母MD2/1(pMI6)分泌的α-淀粉酶多约四倍，这表明乳酸克鲁维酵母是重组蛋白分泌和生产的有效宿主细胞。
