Cinnamaldehyde (CA), a bioactive compound from cinnamon, exhibits diverse pharmacological activities including anti-inflammatory and anticancer effects. However, comprehensive exploration of the target landscape of CA at site-specific resolution remains challenging. Herein, we developed MOSCAT (MethOxyamine-enabled Site-specific Cinnamaldehyde Tagging), a probe-free chemical proteomic strategy for mapping cinnamaldehyde-targeted proteins in living cells. Unlike existing probe-based approaches requiring synthetic CA derivatives, MOSCAT directly captures native CA-protein adducts by exploiting the intrinsic aldehyde functionality of CA, providing unbiased target profiling with residue-level resolution. Using MOSCAT, we identified 632 CA-modification sites across 480 proteins in human cells. Remarkably, over 70% of these sites overlap with other post-translational modifications, particularly S-nitrosylation and S-sulfenylation, revealing mechanistic links to the anti-inflammatory activity of CA. Notably, we discovered that CA covalently modifies Cys93 of GPX4, a conserved residue critical for ferroptosis regulation. This modification triggers proteasome-mediated GPX4 degradation, identifying a specific covalent engagement site associated with CA-induced ferroptosis. Our findings demonstrate MOSCAT as a powerful platform for elucidating molecular mechanisms of electrophilic natural products and highlight GPX4 Cys93 as a promising druggable site for CA-based therapeutic interventions.
Tian et al. (Wed,) studied this question.