Identification of Proteins Containing Cysteine Residues That Are Sensitive to Oxidation by Hydrogen Peroxide at Neutral pH

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Title
Identification of Proteins Containing Cysteine Residues That Are Sensitive to Oxidation by Hydrogen Peroxide at Neutral pH
Author(s)
Jae Ryong Kim; Hae Won Yoon; Ki Sun Kwon; Seung Rock Lee; Sue Goo Rhee
Bibliographic Citation
Analytical Biochemistry, vol. 283, no. 2, pp. 214-221
Publication Year
2000
Abstract
A procedure for detecting proteins that contain H2O2-sensitive cysteine (or selenocysteine) residues was developed as a means with which to study protein oxidation by H2O2 in cells. The procedure is based on the facts that H2O2 and biotin-conjugated iodoacetamide (BIAM) selectively and competitively react with cysteine residues that exhibit a low pK(a), and that the decrease in the labeling of cell lysate proteins with BIAM caused by prior exposure of cells to H2O2 or to an agent that induces H2O2 production can be monitored by streptavidin blot analysis. This procedure was applied to rat pheochromocytoma PC12 cells directly treated with H2O2, mouse hippocampal HT22 cells in which H2O2 production was induced by glutamate, and human erythroleukemia K562 cells in which H2O2 production was induced by phorbol myristate acetate. It revealed that several cell proteins contain cysteine or selenocysteine residues that are selectively oxidized by H2O2. Three of these H2O2-sensitive proteins were identified as a member of the protein disulfide isomerase family, thioredoxin reductase, and creatine kinase, all of which were previously known to contain at least one reactive cysteine or selenocysteine at their catalytic sites. This procedure should thus prove useful for the identification of proteins that are oxidized by H2O2 generated in response to a variety of extracellular agents.
Keyword
assay of protein cysteine oxidationhydrogen peroxideprotein oxidationselenocysteine oxidation
ISSN
0003-2697
Publisher
Elsevier
DOI
http://dx.doi.org/10.1006/abio.2000.4623
Type
Article
Appears in Collections:
Aging Convergence Research Center > 1. Journal Articles
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