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bioRxiv · 10.64898/2026.05.05.721470

Analysis of Confounding Factors in Reactive Cysteine Profiling Reveals Enhanced Chromatin-Protein Association via CDK7 Inhibition by THZ1

Yang, K.·Li, S.·Li, B.·Richards, D.·Dong, K.·Seneviratne, U.·Lee, W.·Iannetta, A.·Xu, H.·Gygi, S.·Yu, Q.
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Abstract

Recent advances in activity-based proteome profiling (ABPP) have enabled global mapping of cysteine ligandability, uncovering novel biological insights and opportunities for identifying disease vulnerabilities. While both live cell-based and native lysate-based ABPP have been applied, how cysteine ligandability differs between these systems and what factors influence these measurements remain unclear. Building on our previous development of a high-throughput TMT-ABPP workflow for native lysates, here we adapt the protocol for live cells and systematically compare cysteine ligandability across both platforms. Our analysis reveals three major contributors to the discrepancies: in-cellulo cysteine accessibility, protein abundance changes, and protein relocalization. Notably, we highlight that CDK7 inhibitor THZ1 induces substantial protein relocalization and promotes chromatin binding. Together, these results provide a practical framework for ABPP experimental design and data interpretation, supporting more accurate application of ABPP in functional proteomics and drug discovery.

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Yang, K., Li, S., Li, B., Richards, D., Dong, K., Seneviratne, U., Lee, W., Iannetta, A., Xu, H., Gygi, S., Yu, Q.. 2026-05-07. Analysis of Confounding Factors in Reactive Cysteine Profiling Reveals Enhanced Chromatin-Protein Association via CDK7 Inhibition by THZ1. https://doi.org/10.64898/2026.05.05.721470

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