Kortemme Tanja, Joachimiak Lukasz A, Bullock Alex N, Schuler Aaron D, Stoddard Barry L, Baker David
Howard Hughes Medical Institute & Department of Biochemistry, Box 357350, University of Washington, Seattle, Washington 98195-7350, USA.
Nat Struct Mol Biol. 2004 Apr;11(4):371-9. doi: 10.1038/nsmb749. Epub 2004 Mar 21.
We developed a 'computational second-site suppressor' strategy to redesign specificity at a protein-protein interface and applied it to create new specifically interacting DNase-inhibitor protein pairs. We demonstrate that the designed switch in specificity holds in in vitro binding and functional assays. We also show that the designed interfaces are specific in the natural functional context in living cells, and present the first high-resolution X-ray crystallographic analysis of a computer-redesigned functional protein-protein interface with altered specificity. The approach should be applicable to the design of interacting protein pairs with novel specificities for delineating and re-engineering protein interaction networks in living cells.
我们开发了一种“计算性的第二位点抑制子”策略，以重新设计蛋白质-蛋白质界面的特异性，并将其应用于创建新的特异性相互作用的脱氧核糖核酸酶抑制剂蛋白对。我们证明，设计的特异性转换在体外结合和功能测定中成立。我们还表明，设计的界面在活细胞的自然功能环境中具有特异性，并展示了对具有改变特异性的计算机重新设计的功能性蛋白质-蛋白质界面的首次高分辨率X射线晶体学分析。该方法应适用于设计具有新特异性的相互作用蛋白对，以描绘和重新构建活细胞中的蛋白质相互作用网络。
