小分子
蛋白质组
结合位点
光亲和标记
计算生物学
化学
生物
生物化学
作者
Jacob M. Wozniak,Weichao Li,Paolo Governa,Liyun Chen,Appaso Mahadev Jadhav,Ashok Dongre,Stefano Forli,Christopher G. Parker
标识
DOI:10.1038/s41589-023-01514-z
摘要
Photoaffinity probes are routinely utilized to identify proteins that interact with small molecules. However, despite this common usage, resolving the specific sites of these interactions remains a challenge. Here we developed a chemoproteomic workflow to determine precise protein binding sites of photoaffinity probes in cells. Deconvolution of features unique to probe-modified peptides, such as their tendency to produce chimeric spectra, facilitated the development of predictive models to confidently determine labeled sites. This yielded an expansive map of small-molecule binding sites on endogenous proteins and enabled the integration with multiplexed quantitation, increasing the throughput and dimensionality of experiments. Finally, using structural information, we characterized diverse binding sites across the proteome, providing direct evidence of their tractability to small molecules. Together, our findings reveal new knowledge for the analysis of photoaffinity probes and provide a robust method for high-resolution mapping of reversible small-molecule interactions en masse in native systems. A chemoproteomic workflow was developed to determine the interaction sites of photoaffinity probes in cells, enabling the identification of diverse binding pockets and providing evidence of their tractability to small-molecule action.
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