Jiang Zheng-Bing, Song Hui-Ting, Gupta Nishith, Ma Li-Xin, Wu Zhen-Bin
College of Life Science, Hubei University, Wuhan 430062, PR China.
Protein Expr Purif. 2007 Nov;56(1):35-9. doi: 10.1016/j.pep.2007.07.003. Epub 2007 Jul 14.
The lipase genes of Yarrowia lipolytica, LIPY7 and LIPY8, fused with FLO-flocculation domain sequence from Saccharomyces cerevisiae at their N-termini, were expressed in Pichia pastoris KM71. Following the induction with methanol, the recombinant proteins were displayed on the cell surface of P. pastoris, as confirmed by the confocal laser scanning microscopy. The LipY7p and LipY8p were anchored on P. pastoris via the flocculation functional domain of Flo1p. The surface-displayed lipases were characterized for their application as the whole-cell biocatalyst. These lipases can also be cleaved off from their anchor by enterokinase treatment to yield functionally active proteins in the supernatant offering an alternative purification method for LipY7p and LipY8p.
解脂耶氏酵母的脂肪酶基因LIPY7和LIPY8在其N端与来自酿酒酵母的FLO絮凝结构域序列融合，在毕赤酵母KM71中表达。用甲醇诱导后，重组蛋白显示在毕赤酵母的细胞表面，共聚焦激光扫描显微镜证实了这一点。LipY7p和LipY8p通过Flo1p的絮凝功能结构域锚定在毕赤酵母上。对表面展示的脂肪酶作为全细胞生物催化剂的应用进行了表征。这些脂肪酶也可以通过肠激酶处理从其锚定物上切割下来，从而在上清液中产生功能活性蛋白，为LipY7p和LipY8p提供了另一种纯化方法。
