Cells exposed to oxidative stress undergo extensive remodeling of protein structure and metabolism through redox-dependent modifications. Among these, cysteine-based disulfide bonds play critical roles but are frequently masked in conventional proteomic workflows that rely on reducing agents. To overcome this limitation, we combined non-reducing tandem mass tag (TMT) proteomics with LC–MS-based metabolomics in H₂O₂-stimulated MDA-MB-231 breast cancer cells. This integrative strategy revealed more than 1,000 proteins containing disulfide crosslinks, identified using the DBond algorithm. The crosslinks were distributed in a highly selective manner, with enrichment at redox-sensitive cysteine sites located near positively charged residues and distinct patterns across homologous isoforms. Parallel metabolomic profiling uncovered pathway-specific bottlenecks, particularly in glycolysis, the tricarboxylic acid cycle, and nucleotide synthesis. Together, these results indicate that disulfide bonds act as finely tuned regulators of protein function and cellular metabolism under oxidative conditions. Our findings demonstrate the unique advantages of non-reducing proteomics for uncovering redox-controlled protein interaction networks and structural transitions.