Construction of artificial higher order protein complexes allows sampling of structural architectures and functional features not accessible by classical monomeric proteins. Here, we combine in silico modelling with expanded genetic code facilitated strain promoted azide-alkyne cycloaddition to construct artificial complexes that are structurally integrated protein dimers and demonstrate functional synergy. Using fluorescent proteins sfGFP and Venus as models, homodimers and heterodimers are constructed that switched ON once assembled and display enhanced spectral properties. Symmetrical crosslinks are found to be important for functional enhancement. The determined molecular structure of one artificial dimer shows that a new long-range polar network comprised mostly of organised water molecules links the two chromophores leading to activation and functional enhancement. Single molecule analysis reveals the dimer is more resistant to photobleaching spending longer times in the ON state. Thus, genetically encoded bioorthogonal chemistry can be used to generate truly integrated artificial protein complexes that enhance function.

人工高阶蛋白质复合物的构建允许采样经典单体蛋白质无法获得的结构结构和功能特征。在这里，我们将计算机模拟与扩展的遗传密码结合起来，以促进菌株促进叠氮化物-炔烃环加成反应，从而构建结构上整合了蛋白质二聚体并显示出功能协同作用的人工复合物。使用荧光蛋白sfGFP和Venus作为模型，构建同二聚体和异二聚体，它们一旦组装就打开，并显示出增强的光谱特性。发现对称的交联对于功能增强很重要。确定的一个人工二聚体的分子结构表明，一个新的远程极性网络主要由有组织的水分子组成，将两个发色团连接在一起，从而导致活化和功能增强。单分子分析表明，二聚体在开态下花费更长的时间对光致漂白具有更强的抵抗力。因此，遗传编码的生物正交化学可用于产生增强功能的真正整合的人工蛋白质复合物。