Proteomic Profiling of Oxidative Stress Response Proteins Using Methionine Sulfoxide-Inspired ABPP Probes

Methionine oxidation is a significant signature of oxidative stress. In nature, oxidative stress from reactive oxygen species or reactive chlorine species (ROS/RCS) serves as a fundamental defense mechanism against pathogenic sources. However, previous tools for studying methionine oxidation have depended on sulfoxide reduction strategies, making them unsuitable for use under oxidative stress conditions. Moreover, no ABPP tools have been reported for studying the activity of cellular methionine sulfoxide reductases. In this paper, we present a cysteine-targeted chemical probe inspired by methionine sulfoxide for profiling oxidative stress response proteins, including methionine sulfoxide reductases. Our probe efficiently labels nucleophilic cysteines of methionine sulfoxide reductases and functions as an activity-based protein profiling tool. The probe demonstrated good reactivity and stability, making it applicable to both in vitro and in vivo experiments under oxidative stress conditions. We showed that our probe labeled oxidative stress-related proteins in E. coli under hypochlorite stress. Finally, we demonstrated that our ABPP probe offers complementary cysteine reactivity profiles compared to conventional probes, efficiently identifying proteins such as DJ-1 superfamily members that are poorly captured by iodoacetamide-alkyne. We believe that our probe can be used to reveal the biological mechanisms by which bacteria sense strong oxidants and endure oxidative stress, such as that encountered with neutrophils.