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根据人胶原蛋白ⅩⅧNC结构域C末端编码内皮细胞抑制生长素 (Endostatin)的成熟多肽序列 ,人工合成了由毕赤酵母 (Pichiapastoris)中偏爱密码子组成的内皮细胞抑制生长素基因序列。该基因以N端融合的方式正确插入毕赤酵母诱导型表达载体pPICZαA中。通过电激将线性化的重组质粒转化到毕赤酵母SMD1 1 68细胞中 ,筛选获得表达具有生物活性的内皮细胞生长抑制素的高产菌株。在摇瓶发酵水平上 ,产量达到 80mg L。而利用生物反应器进行高密度发酵 ,Endostatin的产量可达 1 2 5mg L。纯化后的Endostatin具有抑制小鼠血管内皮细胞增殖的活性
Based on the mature polypeptide sequence encoded by the endostatin of human collagen XVIIINC domain at the C-terminus, the endothelial cell inhibitory auxin gene sequence composed of the preferred codons in Pichia pastoris was artificially synthesized. The gene was inserted into Pichia pastoris-inducible expression vector pPICZαA by N-terminal fusion. The linearized recombinant plasmids were transformed into Pichia pastoris SMD1 1 68 cells by electroporation and screened to obtain high-yielding strains expressing bioactive endostatin. In the shake flask fermentation level, the output reached 80mg L. The use of bioreactors for high-density fermentation, endostatin production up to 125mg. Purified Endostatin has the activity of inhibiting the proliferation of mouse vascular endothelial cells