The present review outlines the most important classes and selected examples of oxidative protein modifications, elucidates the chemistry beyond their formation and discusses available methods for detection and analysis. Moreover, progress in the development of detection and quantification methods facilitates analysis time and effort and contributes to their future applicability in clinical routine. Certain protein oxidation products are chemically stable and formed in large quantity, which makes them promising candidates to become biomarkers of oxidative damage. A rising number of publications provide evidence for their involvement in the onset and progression of diseases as well as aging processes. Reversible modifications are relevant in physiological processes and constitute signaling mechanisms (“redox signaling”), while non-reversible modifications may contribute to pathological situations and several diseases. Consequently, a wide variety of distinct posttranslational protein modifications is formed by protein oxidation, glycoxidation, and lipoxidation. Additionally, oxidative stress is able to degrade lipids and carbohydrates to highly reactive intermediates, which eventually attack proteins at various functional sites. Proteins are major targets for oxidation reactions, because of their rapid reaction rates with oxidants and their high abundance in cells, extracellular tissues, and body fluids. This level of sensitivity allowed measurement of protein carbonylation in individual Drosophila.Generation of reactive oxygen species and related oxidants is an inevitable consequence of life. The detection limit is 0.19☐.04pmol of carbonyl, and 60ng of protein is sufficient to measure protein carbonyl content. It also readily dissolves 2,4-dinitrophenylhydrazine and wets polyvinylidene difluoride (PVDF) membranes. Dimethyl sulfoxide was employed as the solvent because it very efficiently extracts proteins from tissues and keeps them soluble. We developed an immunochemical dot blot method for quantitation of protein carbonylation in homogenates or purified proteins. It is frequently measured spectrophotometrically or immunochemically by derivatizing proteins with the classical carbonyl reagent, 2,4-dinitrophenylhydrazine. 241-245 ISSN: 0003-2697 Subject: Drosophila, detection limit, dimethyl sulfoxide, proteins, solvents, tissues Abstract: Protein carbonylation is the most commonly used measure of oxidative modification of proteins. Levine Source: Analytical biochemistry 2012 v.423 no.2 pp. Quantitation of protein carbonylation by dot blot Author: Nancy B.
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