Protein folding stabilities are a major determinant of oxidation rates for buried methionine residues

Abstract

The oxidation of protein-bound methionines to form methionine sulfoxides has a broad range of biological ramifications, making it important to delineate factors that influence methionine oxidation rates within a protein. This is especially important for biopharmaceuticals, where oxidation can lead to deactivation and degradation. Previously, neighboring residue effects and solvent accessibility (SA) have been shown to impact the susceptibility of methionine residues to oxidation. In this study, we provide proteome-wide evidence that oxidation rates of buried methionine residues are also strongly influenced by the thermodynamic folding stability of proteins. We surveyed the E. coli proteome using several proteomic methodologies and globally measured oxidation rates of methionines in the presence and absence of tertiary structure, as well as folding stabilities of methionine containing domains. The data indicate that buried methionines have a wide range of protection factors against oxidation which correlate strongly with folding stabilities. Concordantly, we show that in comparison to E. coli, the proteome of the thermophile T. thermophilus is significantly more stable and thus more resistant to methionine oxidation. These results indicate that oxidation rates of buried methionines from the native state of proteins can be used as a metric of folding stability. To demonstrate the utility of this correlation, we used native methionine oxidation rates to survey the folding stabilities of E. coli and T. thermophilus proteomes at various temperatures and suggest a model that relates the temperature dependence of the folding stabilities of these two species to their optimal growth temperatures.

Folding stabilities of ribosome-bound nascent polypeptides probed by mass spectrometry

“…Since the kinetics of protein folding are generally faster than oxidation rates, the folding reaction is a pre-equilibrium that modulates the oxidation rate. Thus, buried methionine residues have a slower rate of oxidation within more stable protein regions 37 .…”

“…By making some simplifying assumptions regarding the nature of the methionine oxidation reaction, the differences in oxidation rates between the ribosome-bound and soluble forms of proteins can be used to approximate the difference in the free folding energy between the two. The details of the kinetic model used for this analysis has been described previously 3, 37 and outlined in the Methods section below. Briefly, if we make the assumptions that 1) rate of folding is greater than the rate of unfolding (i.e.…”

“…SASA measurements were computed for E. coli available proteins as previously described 37 . Briefly, solvent accessible surface areas of methionine residues were calculated in Pymol, with dot density set to 3.…”

“…Methionine is a readily oxidizable amino acid and can be converted to methionine sulfoxide by the oxidizing agent hydrogen peroxide (H2O2) 33 . The oxidation rate of a methionine residue within a protein is contingent on its local structural environment [34][35][36][37] . For a methionine that is fully buried in the protein core, unfolding is required for oxidation.…”

Methionine Alkylation as an Approach to Quantify Methionine Oxidation Using Mass Spectrometry

“…Methionine is a sulfur-containing amino acid that is susceptible to oxidation. 1,2 Sidechain oxidation converts nonpolar methionine residues to polar methionine sulfoxides, and this change in hydrophobicity can dramatically alter the structure and function of proteins. [3][4][5][6][7] The conversion of methionine to methionine sulfoxide can occur chemically through reactions with reactive oxygen species (ROS), or enzymatically through the action of specific oxygenases.…”