Novel biomarker pipeline to probe the oxidation sites and oxidation degrees of hemoglobin in bovine erythrocytes exposed to oxidative stress.
Research on biomarkers for protein oxidation might give insight into the mechanistic mode of oxidative stress. In the work present here, a novel pipeline was established to probe the oxidation mechanism of bovine hemoglobin (Hb) with its oxidation products serving as the biomarkers. Reactive oxygen species generated by irradiation were used to mimic oxidative stress conditions to oxidize Hb in bovine erythrocytes. After Hb extraction and digestion, oxidized peptides in the tryptic fragments were assigned by comparison with the extracted ion chromatography spectra of native peptide from the control sample. Subsequent tandem mass spectrometry analysis of these peptides proved that oxidation was limited to partially exposed amino acid residues (α-Phe36 , β-Met1 , β-Trp14 , for instance) in Hb. Quantitation analysis on these oxidized peptides showed that oxidation degrees of target sites had positive correlations with the extended oxidation dose and the oxidation processes were also controlled by residues types. Compared with the conventional protein carbonyl assay, the identified oxidized products were feasibility biomarkers for Hb oxidation, indicating that the proposed biomarker pipeline was suitable to provide specific and valid information for protein oxidation. Copyright © 2015 John Wiley & Sons, Ltd.
College of Population, Resources and Environment, Shandong Normal University, Jinan, 250014, People's Republic of China.
Wansong
W
Zong
College of Population, Resources and Environment, Shandong Normal University, Jinan, 250014, People's Republic of China.
Xiaoning
X
Wang
College of Population, Resources and Environment, Shandong Normal University, Jinan, 250014, People's Republic of China.
Chuanxi
C
Yang
College of Population, Resources and Environment, Shandong Normal University, Jinan, 250014, People's Republic of China.
Yonggang
Y
Du
College of Population, Resources and Environment, Shandong Normal University, Jinan, 250014, People's Republic of China.
Weijun
W
Sun
College of Population, Resources and Environment, Shandong Normal University, Jinan, 250014, People's Republic of China.
Zhenzhen
Z
Xu
D000595:Amino Acid Sequence; D000818:Animals; D015415:Biomarkers / Q000378:metabolism*; D002417:Cattle; D002853:Chromatography, Liquid / Q000379:methods; D004912:Erythrocytes / Q000378:metabolism*; D006454:Hemoglobins / Q000737:chemistry / Q000378:metabolism*; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D017386:Sequence Homology, Amino Acid; D053719:Tandem Mass Spectrometry / Q000379:methods
D015415:Biomarkers; D006454:Hemoglobins
Mass spectrometry; biomarker; hemoglobin; liquid chromatography; oxidative stress; protein oxidation