Abstract 1529 Effect of N-Terminal Modifications on MSR Activity

Methionine sidechains can become oxidized by reacting with reactive oxygen species (ROS), resulting in the formation of methionine sulfoxides (MetO).This oxidative damage can be reversed by a class of enzymes referred to as methionine sulfoxide reductases (MSRs).Methionines also play an important role in translation as initiator residues of newly synthesized proteins.In prokaryotes and the mitochondria and chloroplasts of eukaryotes, initiator methionines are formylated at the N-terminal (fMet), Our laboratory has previously shown that oxidation of initiator methionines leads to mis-processing of nascent polypeptides and formylation of initiator methionines enhances their reduction and repair by MSRs.However, the mechanism by which formylation of MetOs enhances their reduction by Msrs remains unknown.To gain insights into the effects of modifications of MetOs on MSR activity, we generated synthetic MetO-containing peptides with alternative N-terminal modifications including methylation, acetylation and propionylation.We then measured the activity of MSR on these modified peptides using in vitro biochemical assays.Our initial results indicate that, similar to formylation, acetylation and propionylation of Ntermini increases MSR efficiency on oxidized peptides.Methylation of the N-termini, however, leads to decreased efficiency of MSRs on oxidized peptides.These results suggest that adding a polar carbonyl group to N-termini of MetOs enhances their interaction with MSRs and increases their rates of reduction.Future experiments with different MSRs can help determine whether these effects are universal across all MSRs and provide further insights into the mechanism of reduction of MetOs by MSRs.