A General Strategy for the Design and Evaluation of Heterobifunctional Tools: Applications to Protein Localization and Phase Separation
A General Strategy for the Design and Evaluation of Heterobifunctional Tools: Applications to Protein Localization and Phase Separation
复制标题
异双功能工具设计和评估的一般策略：在蛋白质定位和相分离中的应用
DOI：
10.1002/cbic.202200209
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发表时间：
2022
期刊：
影响因子：
3.2
通讯作者：
Chenoweth, David M.
中科院分区：
文献类型：
--
作者：
Lackner, Rachel M.;O'Connell, Will;Zhang, Huaiying;Chenoweth, David M.
为了模拟自然界中存在的时空控制水平，使用化学诱导二聚化（CID）工具来操纵蛋白质-蛋白质相互作用。虽然已知连接体组成影响异双功能化合物的速度和效率，但建模或体外实验通常不足以预测最佳连接体结构。 这可以归因于三元复合物形成的复杂性和影响细胞内探针有效浓度的重叠因素，例如外排和被动渗透性。在这里，我们合成了一个具有不同连接结构的模块化化学工具库，并在活细胞中进行定量显微镜检查，以真实的实时可视化二聚化。我们使用我们优化的探针来证明我们将感兴趣的蛋白质（POI）招募到线粒体、细胞膜和细胞核的能力。最后，我们诱导和监测本地和全球相分离。我们强调了定量方法的重要性，动态系统的连接器优化，并介绍了新的，综合可访问的工具，快速控制蛋白质定位。
To mimic the levels of spatiotemporal control that exist in nature, tools for chemically induced dimerization (CID) are employed to manipulate protein‐protein interactions. Although linker composition is known to influence speed and efficiency of heterobifunctional compounds, modeling orin vitroexperiments are often insufficient to predict optimal linker structure. This can be attributed to the complexity of ternary complex formation and the overlapping factors that impact the effective concentration of probe within the cell, such as efflux and passive permeability. Herein, we synthesize a library of modular chemical tools with varying linker structures and perform quantitative microscopy in live cells to visualize dimerization in real‐time. We use our optimized probe to demonstrate our ability to recruit a protein of interest (POI) to the mitochondria, cell membrane, and nucleus. Finally, we induce and monitor local and global phase separation. We highlight the importance of quantitative approaches to linker optimization for dynamic systems and introduce new, synthetically accessible tools for the rapid control of protein localization.