Isolation and Characterization of Oxidized Lysozyme Variants Produced by a Copper (II)/Hydrogen Peroxide Metal-Catalyzed Oxidation System

Abstract

Protein oxidation has been correlated with several chronic diseases including Alzheimer's disease, Parkinson's disease, and cataractogenesis. The purpose of this project was to isolate and characterize the various oxidized forms of hen egg white lysozyme that were produced by a copper(II)/hydrogen peroxide metal-catalyzed oxidation system. Five oxidized protein variants were purified using high performance liquid chromatography on a cation-exchange column. Tandem mass spectrometry determined that several amino acids were oxidized in each variant with histidine 15 being the most readily oxidized residue. Bacteriolytic assays showed decreased activity of Peaks IB, IIB, and III (31.4%, 61.2%, and 86.5%, respectively) relative to native enzyme while the activity for Peaks IV and V was greater than that of native enzyme (215% and 308%, respectively). Crystals of Peaks IB, III, IV, and V were grown, but attempts to determine the crystal structure were unsuccessful.