利用BglBrick方法进行HSA/Exendin-4长效融合蛋白的快速组装

Rapid Assembly of Long-term HSA /Exendin 4 Fusion Protein via BglBrick Method

目的:通过合成生物学的BglBrick方法快速构建不同种类HSA/Exendin-4长效融合蛋白,为进行各HSA/Exendin-4融合蛋白生物学活性的筛选比较奠定基础。方法:选用酵母表达载体pPICZαA质粒,构建pPICZαA-E4和pPICZαA-HSA两种基础Brick表达载体,运用BglBrick方法,实现了不同数目Exendin-4分子在HSA的不同末端的快速组装。将构建出的10种HSA/Exendin-4融合蛋白,转入毕赤酵母菌KM71H中,用甲醇诱导目的蛋白的表达。结果:用BglBrick方法构建的HSA/Exendin-4融合蛋白在毕赤酵母中都成功诱导表达出相应的目的蛋白。结论:利用BglBrick方法能够实现HSA长效融合蛋白的快速组装。

Objective： Different kinds of HSA/Exendin4 fusion protein are rapidly constructed using BglBrick method of synthetic biology. It lays a foundation for screening and comparison of the bioactivity of fusion protein in each HSA/Exendin-4. Method： Two basic Brick expression vectors, pPICZαA-Exendin4 and pPTCZαA-HSA, are constructed based on the yeast expression vector, pPICZαA plasmid. Rapid assemblies of different amounts of Exendin4 molecules at the terminals of HSA are achieved via BglBrick method. Ten constructed HSA/Exendin-4 fusion proteins are integrated into the chromosome of Pichia pastoris KM71H, and the corresponding target proteins are expressed after methanol induction. Result：The constructed HSA/Exendin-4 fusion proteins via BglBrick method successfully express the corresponding target protein in Pichia pastoris after methanol induction. Conclusion： The use of BglBrick method can help fulfill the rapid assembly of long-term HAS fusion proteins.