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

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。

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

Objective: Different kinds of HSA/Exendin-4 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-Exendin-4 and pPICZαA-HSA, are constructed based on the yeast expression vector, pPICZαA plasmid. Rapid assemblies of different amounts of Exendin-4 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.