Understanding Oxidative Instability of Protein Pharmaceuticals

Abstract

Mechanism of oxidation of methionine residues in protein pharmaceuticals by hydrogen peroxide was investigated via ab initio calculations. Specifically, two reactions, hydrogen transfer of hydrogen peroxide to form water oxide and the oxidation of dimethyl sulfide (DMS) by hydrogen peroxide to form dimethyl sulfoxide, were studied as models of these processes in general. Solvent effects are included both via including explicitly water molecules and via the polarizable continuum model. Specific interactions including hydrogen bonding with 2-3 water molecules can provide enough stabilization for the charge separation at the activation complex. The major reaction coordinates of the reaction are the breaking of the O-O bond of H₂O₂ and the formation of the S-O bond, the transfer of hydrogen to the distal oxygen of hydrogen peroxide occurring after the system has passed the transition state. Reaction barriers of the hydrogen transfer of H₂O₂ are in average of 10 kcal/mol or higher than the oxidation of DMS. Therefore, a two step oxidation mechanism in which the transfer of hydrogen atom occurs first to form water oxide and the transfer of oxygen to substrate occurs as the second step, is unlikely to be correct. Our proposed oxidation mechanism does not suggest pH dependence of oxidation rate within a moderate range around neutral pH (i.e. under conditions in which hydronium and hydroxide ions do not participate directly in the reaction), and it agrees with experimental observations over moderate pH values.