title: Inactivation of human peroxiredoxin I during catalysis as the result of the oxidation of the catalytic site cysteine to cysteine-sulfinic acid.
abstract: By following peroxiredoxin I (Prx I)-dependent NADPH oxidation spectrophotometrically, we observed that Prx I activity decreased gradually with time. The decay in activity was coincident with the conversion of Prx I to a more acidic species as assessed by two-dimensional gel electrophoresis. Mass spectral analysis and studies with Cys mutants determined that this shift in pI was due to selective oxidation of the catalytic site Cys(51)-SH to Cys(51)-SO(2)H. Thus, Cys(51)-SOH generated as an intermediate during catalysis appeared to undergo occasional further oxidation to Cys(51)-SO(2)H, which cannot be reversed by thioredoxin. The presence of H(2)O(2) alone was not sufficient to cause oxidation of Cys(51) to Cys(51)-SO(2)H. Rather, the presence of complete catalytic components (H(2)O(2), thioredoxin, thioredoxin reductase, and NADPH) was necessary, indicating that such hyperoxidation occurs only when Prx I is engaged in the catalytic cycle. Likewise, hyperoxidation of Cys(172)/Ser(172) mutant Prx I required not only H(2)O(2), but also a catalysis-supporting thiol (dithiothreitol). Kinetic analysis of Prx I inactivation in the presence of a low steady-state level (<1 microm) of H(2)O(2) indicated that Prx I was hyperoxidized at a rate of 0.072% per turnover at 30 degrees C. Hyperoxidation of Prx I was also detected in HeLa cells treated with H(2)O(2).
authors.affiliation: Laboratory of Cell Signaling, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
authors.forename: Kap-Seok
authors.initials: KS
authors.lastname: Yang
authors.forename: Sang Won
authors.initials: SW
authors.lastname: Kang
authors.forename: Hyun Ae
authors.initials: HA
authors.lastname: Woo
authors.forename: Sung Chul
authors.initials: SC
authors.lastname: Hwang
authors.forename: Ho Zoon
authors.initials: HZ
authors.lastname: Chae
authors.forename: Kanghwa
authors.initials: K
authors.lastname: Kim
authors.forename: Sue Goo
authors.initials: SG
authors.lastname: Rhee
mesh_terms: D000595:Amino Acid Sequence; D000975:Antioxidants / Q000737:chemistry / Q000378:metabolism; D002384:Catalysis; D020134:Catalytic Domain; D003545:Cysteine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D009154:Mutation; D009249:NADP / Q000378:metabolism*; D016877:Oxidants / Q000378:metabolism; D010084:Oxidation-Reduction; D010455:Peptides / Q000378:metabolism; D010544:Peroxidases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D054464:Peroxiredoxins; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D021241:Spectrometry, Mass, Electrospray Ionization; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013880:Thioredoxin-Disulfide Reductase / Q000378:metabolism; D013879:Thioredoxins / Q000378:metabolism
chemical_list: D000975:Antioxidants; D016877:Oxidants; D010455:Peptides; D011994:Recombinant Proteins; D013879:Thioredoxins; D009249:NADP; D006861:Hydrogen Peroxide; D010544:Peroxidases; D054464:Peroxiredoxins; D013880:Thioredoxin-Disulfide Reductase; D003545:Cysteine