PDF(Pubmed)
Abstract：
Protein-protein interactions (PPIs) are central in cell metabolism but research tools for the structural and functional characterization of these PPIs are often missing. Here we introduce broadly applicable immunization (Cross-link PPIs and immunize llamas, ChILL) and selection strategies (Display and co-selection, DisCO) for the discovery of diverse nanobodies that either stabilize or disrupt PPIs in a single experiment. We apply ChILL and DisCO to identify competitive, connective, or fully allosteric nanobodies that inhibit or facilitate the formation of the SOS1•RAS complex and modulate the nucleotide exchange rate on this pivotal GTPase in vitro as well as RAS signalling in cellulo. One of these connective nanobodies fills a cavity that was previously identified as the binding pocket for a series of therapeutic lead compounds. The long complementarity-determining region (CDR3) that penetrates this binding pocket serves as pharmacophore for extending the repertoire of potential leads.
摘要：
蛋白质-蛋白质相互作用（PPIs）是细胞代谢的核心，但经常缺少用于表征这些PPIs的结构和功能的研究工具。在这里，我们介绍了广泛适用的免疫(交联PPI和免疫美洲驼，冷静)和选择策略(显示和共同选择，DisCO），用于发现在单个实验中稳定或破坏PPI的多种纳米抗体。我们应用ChILL和DisCO来识别有竞争力的，结缔组织，或完全变构的纳米抗体，其抑制或促进SOS1•RAS复合物的形成，并在体外调节该关键GTP酶的核苷酸交换速率以及在纤维素酶中的RAS信号传导。这些连接纳米体中的一个填充了先前被鉴定为一系列治疗性先导化合物的结合袋的腔。穿透该结合袋的长互补决定区(CDR3)用作药效团,用于延伸潜在前导的库。
