抑肽酶基因的克隆和表达

抑肽酶作为天然非特异性丝氨酸蛋白酶抑制剂，用途广泛。目前抑肽酶制品主要从牛肺中提取。用基因重组技术，将抑肽酶结构基因导入表达载体pGrxA，并在抑肽酶基因上游引入FXa识别位点。将重组质粒转化至Origami^TM中，用异丙基硫代β-D半乳糖苷(IPTG)诱导表达目的蛋白，产物以可溶性形式存在于胞内。将菌体超声破壁和离心，融合蛋白经分子筛层析和离子交换层析纯化后，用FXa切割该融合蛋白可得到N端不含多余氨基酸残基且与天然构像一致的活性抑肽酶。

Cloning and Expression of Recombinant Bovine Pancreatic Trypsin Inhibitor in E.coli.

Aprotinin, also known as bovine pancreatic trypsin inhibitor(BPTI), is a serine proteinase inhibitor. It is widely used for clinical purpose. Aprotinin is naturally extracted from bovine lungs. In this paper, production of aprotinin with genetic engineering method was described. The recombinant gene for BPTI with FXa cleavage site was fused into expression vector pGrxA. When induced with IPTG, the fusion protein was expressed in E.coli. Origami TM up to 10% of total protein. After purification, the fusion protein was specifically cleaved with FXa and the recombinant BPTI without N-terminal extended amino acid residual was obtained, and the recombinant BPTI appears the same characterization as native BPTI.