Human Mn-superoxide dismutase acetylation protects from enzyme nitration and inactivation

Manganese superoxide dismutase (MnSOD) is a critical enzyme responsible for detoxifying superoxide radicals in mitochondria, thereby ensuring oxidative balance within cells. Post-translational modifications (PTMs), such as acetylation and nitration, significantly influence MnSOD's catalytic efficiency. Site-specific nitration of MnSOD at tyrosine 34 by peroxynitrite leads to an irreversible inactivation, which has been widely observed in diverse pathologies. On the other hand, acetylation of MnSOD is a reversible modification that modulates the activity of the enzyme and it is finely regulated by the action of the protein Sirt3, responsible for the deacetylation of a wide variety of mitochondrial enzymes. This study focuses on Lys29 acetylation and its impact on the enzyme's activity and its interplay with peroxynitrite-mediated nitration of Tyr34. Through molecular dynamics (MD) simulations, we observed that acetylation of Lys29 partially obstructs the access channel to the active site, reducing superoxide accessibility. Electrostatic potential calculations further revealed that Lys29 acetylation diminishes the positive charge around the active site, contributing to decreased affinity for superoxide radicals. Brownian dynamics (BD) simulations confirmed a 50 % reduction in the enzyme's association rate constant (kon) for superoxide upon Lys29 acetylation. In contrast, Lys98 acetylation had a minor effect on kon. In vitro studies also supported our findings and showed that acetylation could play a role in the irreversible inactivation of MnSOD by peroxynitrite, likely by sterically hindering Tyr34 nitration. These findings highlight the role of acetylation as a reversible protective mechanism that can regulate superoxide and peroxynitrite accessibility to MnSOD under stress conditions.