Using alchemical free energy simulations, the study shows cysteine oxidation can either destabilize or stabilize proteins depending on context—predicting decreased stability and local unfolding for URN1-FF while oxidation of DJ-1 increases monomer stability and slightly destabilizes dimerization.

Because DJ-1 is a Parkinson’s-linked, redox-sensitive protein, the mechanistic insights and general computational protocol could inform strategies to modulate DJ-1 stability or redox state for neuroprotective therapeutic development.

The extracellular environment but also cellular metabolism can generate oxidative stress that can chemically modify and damage protein molecules. The sulfur containing amino acid cysteine (CYS) is particularly vulnerable to oxidation. The molecular details of how CYS oxidation can modulate stability and binding of proteins is still not well understood. Using alchemical free energy simulations, we calculate the change in protein stability and association upon CYS oxidation to different oxidation states for two example proteins. In the case of the URN1 splicing factor FF domain (URN1-FF) the simulations predict a significant decrease in stability upon oxidation in agreement with experiment and the effect also depends on the final oxidation state. In addition, the oxidation leads to conformational changes and partial unfolding at the protein C-terminus. In contrast, for the second system, Parkinson disease protein 7 (DJ-1), CYS oxidation enhances significantly the protein monomer stability again in agreement with the experimental observation and slightly destabilizes homo dimerization. Analysis of the molecular details associated with CYS oxidation in the folded proteins allows us to gain insights into why both stabilizing as well as destabilizing effects can be observed. The CYS oxidation simulation methodology could also serve as a general protocol to analyze single and multiple CYS oxidations in other protein systems and its influence on protein binding and stability.