BIOLOGICAL CONSEQUENCES OF PROTEIN OXIDATION

点击翻译按钮获取中文摘要

Proteins can become oxidized under a variety of conditions in vitro and in vivo. Specific polypeptide changes include formation of aldehyde (carbonyl) groups, generation of methionine sulfoxide and dityrosine, rearrangement of disulfide bonds, cleavage of peptide bonds, and protein aggregation. The consequences of protein oxidative modifications are varied. Some modifications lead to differential susceptibility of a protein to proteolytic cleavage while others cause gain or loss of biological activity. For example, we have found that oxidative modification of fibrinogen inhibits ability of the protein to undergo clotting. Conversely, oxidation of plasma protease inhibitors can lead to an increase of clotting activity. Our research involves examination of the conditions under which proteins acquire oxidative modifications and development of assays that provide a correct assessment of the types of modifications incurred. We determined that the carbohydrate moieties of glycoproteins do not undergo oxidative modification to carbonyl groups under conditions that oxidize amino acid side chains. In addition, we validated the use of a Western blot assay for protein carbonyls for use on glycoproteins. Current studies are aimed at developing an appropriate in vitro model for studying protein oxidative modification in cells exposed to varying growth conditions, including oxidative stress. Because cellular lipids are also targets for oxidative attack and can modify proteins through conjugation reactions, it is important to distinguish between lipid-derived protein modifications and direct protein oxidative modification. We have chosen to work with lymphoma cells that express different levels of the oncogene bcl-2 because resistance to oxidative processes is thought to correlate with expression of high levels of the Bcl-2 protein. The biological consequences of oxidative reactions will be explored by comparing the effects of H2O2 treatment on cells that are either sensitive or resistant to this oxidant.