Department of Biochemistry, University of Washington, Seattle, WA, USA.
Institute for Protein Design, University of Washington, Seattle, WA, USA.
Nat Struct Mol Biol. 2019 Jun;26(6):407-414. doi: 10.1038/s41594-019-0224-z. Epub 2019 May 13.
To discriminate between closely related members of a protein family that differ at a limited number of spatially distant positions is a challenge for drug discovery. We describe a combined computational design and experimental selection approach for generating binders targeting functional sites with large, shape complementary interfaces to read out subtle sequence differences for subtype-specific antagonism. Repeat proteins are computationally docked against a functionally relevant region of the target protein surface that varies in the different subtypes, and the interface sequences are optimized for affinity and specificity first computationally and then experimentally. We used this approach to generate a series of human Frizzled (Fz) subtype-selective antagonists with extensive shape complementary interaction surfaces considerably larger than those of repeat proteins selected from random libraries. In vivo administration revealed that Wnt-dependent pericentral liver gene expression involves multiple Fz subtypes, while maintenance of the intestinal crypt stem cell compartment involves only a limited subset.
区分在空间上相隔有限位置的蛋白质家族中的密切相关成员，是药物发现的一个挑战。我们描述了一种结合计算设计和实验选择的方法，用于生成针对具有大的形状互补界面的功能位点的结合物，以读出亚型特异性拮抗作用的细微序列差异。重复蛋白被计算对接在目标蛋白表面的功能相关区域，该区域在不同的亚型中发生变化，然后首先通过计算，然后通过实验来优化界面序列的亲和力和特异性。我们使用这种方法生成了一系列人卷曲（Fz）亚型选择性拮抗剂，它们具有广泛的形状互补相互作用表面，比从随机文库中选择的重复蛋白大得多。体内给药表明，Wnt 依赖性中央肝基因表达涉及多个 Fz 亚型，而肠隐窝干细胞区室的维持仅涉及有限的亚组。
