Hua Yi, Tay Nicholas E S, Ye Xuanjia, Owen Jeremy A, Liu Hengyuan, Thompson Robert E, Muir Tom W
Department of Chemistry, Princeton University, Princeton, NJ, USA.
Science. 2025 Apr 4;388(6742):68-74. doi: 10.1126/science.adq8540. Epub 2025 Apr 3.
Protein engineering through the ligation of polypeptide fragments has proven enormously powerful for studying biochemical processes. In general, this strategy necessitates a final protein-folding step, constraining the types of systems amenable to the approach. Here, we report a method that allows internal regions of target proteins to be replaced in a single operation. Conceptually, our system is analogous to a DNA transposition reaction but uses orthogonal pairs of engineered split inteins to mediate the editing process. This "protein transposition" reaction is applied to several systems, including folded protein complexes, allowing the efficient introduction of a variety of noncoded elements. By carrying out a molecular "cut and paste" under native protein-folding conditions, our approach substantially expands the scope of protein semisynthesis.
通过连接多肽片段进行蛋白质工程已被证明在研究生化过程方面具有巨大的威力。一般来说，这种策略需要一个最终的蛋白质折叠步骤，限制了适用于该方法的系统类型。在这里，我们报告了一种方法，该方法允许在一次操作中替换目标蛋白质的内部区域。从概念上讲，我们的系统类似于DNA转座反应，但使用经过工程改造的分裂内含肽的正交对来介导编辑过程。这种“蛋白质转座”反应应用于多个系统，包括折叠的蛋白质复合物，从而能够高效引入各种非编码元件。通过在天然蛋白质折叠条件下进行分子“剪切和粘贴”，我们的方法大大扩展了蛋白质半合成的范围。
