Development of light-responsive protein binding in the monobody non-immunoglobulin scaffold
Development of light-responsive protein binding in the monobody non-immunoglobulin scaffold
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DOI：
10.1038/s41467-020-17837-7
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发表时间：
2020-08-13
影响因子：
16.6
通讯作者：
Avalos, Jose L.
中科院分区：
文献类型：
--
作者：
Carrasco-Lopez, Cesar;Zhao, Evan M.;Avalos, Jose L.
单抗体是合成的非免疫球蛋白可定制的蛋白质结合剂，对基础和应用研究非常宝贵，并且作为未来的治疗和诊断工具具有相当大的潜力。根据需要可逆地控制它们与其靶标的结合活性的能力将显著扩大它们在生物技术、医学和研究中的应用。在这里，我们提出，作为原理证明，开发的光控单体（OptoMB），在体外和细胞中工作，其对SH 2结构域目标的亲和力在光照下表现出330倍的结合亲和力的变化。我们证明我们的alpha SH 2-OptoMB可用于直接从粗大肠杆菌中纯化SH 2标记的蛋白质。大肠杆菌提取物，在单一纯化步骤中达到99.8%的纯度和超过40%的收率。凭借其被设计为结合任何感兴趣的蛋白质的能力，OptoMBs有可能找到新的强大的应用程序，作为光可切换的结合剂的未标记的蛋白质与光提供的时间和空间精度。
Monobodies are synthetic non-immunoglobulin customizable protein binders invaluable to basic and applied research, and of considerable potential as future therapeutics and diagnostic tools. The ability to reversibly control their binding activity to their targets on demand would significantly expand their applications in biotechnology, medicine, and research. Here we present, as proof-of-principle, the development of a light-controlled monobody (OptoMB) that works in vitro and in cells and whose affinity for its SH2-domain target exhibits a 330-fold shift in binding affinity upon illumination. We demonstrate that our alpha SH2-OptoMB can be used to purify SH2-tagged proteins directly from crude E. coli extract, achieving 99.8% purity and over 40% yield in a single purification step. By virtue of their ability to be designed to bind any protein of interest, OptoMBs have the potential to find new powerful applications as light-switchable binders of untagged proteins with the temporal and spatial precision afforded by light.