O2-Coupled Copper-Cysteine Redox Chemistry Drives Oxidative Modifications and Higher-Order Assembly of Monomeric Transthyretin

Abstract

Copper-cysteine (Cu-Cys) coordination is pervasive in biology, yet how Cu-thiol redox chemistry engages O2 and translates into protein-level consequences remains incompletely understood. Here, we establish a generalizable protein-based platform that directly captures O2-coupled Cu(II)-Cys redox processes and delineates their structural and functional outcomes. Integrated spectroscopic, mass spectrometric, and computational investigations reveal that Cu(II) coordination to a protein Cys residue initiates redox cycling that generates thiyl radicals and/or disulfide radical anions. This chemically defined process drives disulfide bond formation together with O2-dependent oxidative transformations, producing site-specific Cys modifications, dityrosine cross-links, and higher-order protein assemblies. Under these conditions, these structural changes abolish the protein's anti-amyloidogenic activity and result in apoptotic responses. By directly linking Cu-thiol redox chemistry to protein oxidation and aggregation, this work establishes a fundamental chemical principle by which redox-active metal ions govern protein fate under aerobic conditions. More broadly, these findings provide a mechanistic framework for understanding how metal-mediated oxidative chemistry contributes to protein misfolding and aggregation in human diseases.