PROTAC-Based Proteomics Strategy Uncovers Arsenic-Binding Proteomes

Manipulating intracellular protein degradation systems provides extraordinary opportunities to find targeting proteins critical for drug discovery, as exemplified by proteolysis-targeting chimeras (PROTACs). Here, using trivalent arsenical-PROTAC (As(III)-PROTAC) probes, we developed an in situ proteolysis strategy to uncover the As-binding proteomes. Combined with proteomic analysis, this strategy identified 135 downregulated proteins in A549 human lung carcinoma cells, providing a candidate list for seeking As-binding proteins. Bioinformatics analysis revealed that they were involved in multiple pathways, such as the tricarboxylic acid cycle, DNA replication, and DNA repair. Karyopherin subunit α-2 (KPNA2) and cyclin-dependent kinase 1 (CDK1) could play core roles in the context of protein interactions. Western blotting confirmed the downregulation of embryonic ectoderm development (EED) and phospholipase A-2-activating (PLAA) proteins after the probe treatment. A cellular thermal shift assay (CETSA) exhibited intracellular binding of iAsIII to EED and PLAA. This work demonstrates the development and application of PROTAC probes for screening of As-binding proteomes and establishes a methodological foundation for comprehending the biological effects induced by arsenicals.