Development of a peptide mapping procedure to identify and quantify methionine oxidation in recombinant human α1-antitrypsin

• Published in: Journal of Chromatography A Vol. 942; no. 1; pp. 133 - 143
• Main Authors: Griffiths, Steven W, Cooney, Charles L
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 2002; Elsevier
• Subjects: Aminoacids, peptides. Hormones. Neuropeptides; Analytical, structural and metabolic biochemistry; Antitrypsin; Biological and medical sciences; Fundamental and applied biological sciences. Psychology; Hydrogen peroxide; Methionine; Peptides; Proteins; Hydrogen peroxide; Peptides; Antitrypsin; Methionine; Human; Phase composition; Molecular structure; Coupled method; Active site; Ion pair; HPLC chromatography; Primary structure; Mobile phase; Electrospray; α1-Antitrypsin; Peptide map; Reversed phase chromatography; Analysis method; Peptide fragment; Sulfur peptide; Oxidation; Recombinant protein; Mass spectrometry; Aminoacid sequence; Quantitative analysis; Structural analysis
• ISSN: 0021-9673
• DOI: 10.1016/S0021-9673(01)01350-4

A peptide mapping procedure was developed to identify and quantify methionine oxidation in recombinant human α1-antitrypsin. Due to the protein’s complex structural biochemistry, chromatographic analysis of methionine containing digest peptides was a significant challenge. However, by using a combination of mass spectrometry, protein engineering, and high-temperature reversed-phase liquid chromatography, we were able to identify methionine residues that are susceptible to oxidation by hydrogen peroxide, and quantify their reactivity. Our results show that five of the protein’s 10 methionine residues are susceptible to oxidation at neutral pH, four of which are localized to the active site region.