Department of Molecular and Cellular Biology, University of Washington, Seattle, WA 98195.
Institute for Protein Design, University of Washington, Seattle, WA 98195.
Proc Natl Acad Sci U S A. 2017 Sep 19;114(38):10125-10130. doi: 10.1073/pnas.1706907114. Epub 2017 Sep 1.
The polycomb repressive complex 2 (PRC2) histone methyltransferase plays a central role in epigenetic regulation in development and in cancer, and hence to interrogate its role in a specific developmental transition, methods are needed for disrupting function of the complex with high temporal and spatial precision. The catalytic and substrate recognition functions of PRC2 are coupled by binding of the N-terminal helix of the Ezh2 methylase to an extended groove on the EED trimethyl lysine binding subunit. Disrupting PRC2 function can in principle be achieved by blocking this single interaction, but there are few approaches for blocking specific protein-protein interactions in living cells and organisms. Here, we describe the computational design of proteins that bind to the EZH2 interaction site on EED with subnanomolar affinity in vitro and form tight and specific complexes with EED in living cells. Induction of the EED binding proteins abolishes H3K27 methylation in human embryonic stem cells (hESCs) and at all but the earliest stage blocks self-renewal, pinpointing the first critical repressive H3K27me3 marks in development.
多梳抑制复合物 2（PRC2）组蛋白甲基转移酶在发育和癌症中的表观遗传调控中发挥核心作用，因此，需要有方法能够在特定的发育转变中，以高时空精度破坏该复合物的功能。PRC2 的催化和底物识别功能通过 Ezh2 甲基转移酶的 N 端螺旋与 EED 三甲基赖氨酸结合亚基上的扩展凹槽结合而偶联在一起。原则上，可以通过阻断这种单一相互作用来破坏 PRC2 的功能，但在活细胞和生物体中阻断特定蛋白质-蛋白质相互作用的方法很少。在这里，我们描述了蛋白质的计算设计，这些蛋白质在体外与 EED 上的 EZH2 相互作用位点以纳摩尔亲和力结合，并在活细胞中与 EED 形成紧密和特异性的复合物。EED 结合蛋白的诱导会在人胚胎干细胞（hESC）中消除 H3K27 甲基化，并且除了最早的阶段之外，还会阻止自我更新，从而确定发育过程中第一个关键的抑制性 H3K27me3 标记。
