Department of Biological Chemistry, David Geffen School of Medicine, UCLA, Los Angeles, California 90095, United States.
Department of Chemistry and Biochemistry, UCLA, Los Angeles, California 90095, United States.
Anal Chem. 2021 Feb 2;93(4):2610-2618. doi: 10.1021/acs.analchem.0c04726. Epub 2021 Jan 20.
Mass-spectrometry-based chemoproteomics has enabled the rapid and proteome-wide discovery of functional and potentially 'druggable' hotspots in proteins. While numerous transformations are now available, chemoproteomic studies still rely overwhelmingly on copper(I)-catalyzed azide-alkyne cycloaddition (CuAAC) or 'click' chemistry. The absence of bio-orthogonal chemistries that are functionally equivalent and complementary to CuAAC for chemoproteomic applications has hindered the development of multiplexed chemoproteomic platforms capable of assaying multiple amino acid side chains in parallel. Here, we identify and optimize Suzuki-Miyaura cross-coupling conditions for activity-based protein profiling and mass-spectrometry-based chemoproteomics, including for target deconvolution and labeling site identification. Uniquely enabled by the observed orthogonality of palladium-catalyzed cross-coupling and CuAAC, we combine both reactions to achieve dual labeling. Multiplexed targeted deconvolution identified the protein targets of bifunctional cysteine- and lysine-reactive probes.
基于质谱的化学蛋白质组学已经能够快速且全面地发现蛋白质中具有功能和潜在“可成药”的热点。虽然现在有许多转化方法，但化学蛋白质组学研究仍然主要依赖于铜（I）催化的叠氮-炔环加成（CuAAC）或“点击”化学。缺乏与 CuAAC 在功能上等效且互补的生物正交化学，这阻碍了能够平行检测多种氨基酸侧链的多重化学蛋白质组学平台的发展。在这里，我们确定并优化了用于基于活性的蛋白质谱分析和基于质谱的化学蛋白质组学的铃木-宫浦偶联条件，包括用于目标分解和标记位置鉴定。钯催化交叉偶联和 CuAAC 的正交性使我们能够独特地实现双重标记。多重靶向分解鉴定了双功能半胱氨酸和赖氨酸反应探针的蛋白质靶标。
