title: Release of free amino acids upon oxidation of peptides and proteins by hydroxyl radicals.
abstract: Hydroxyl radical-induced oxidation of proteins and peptides can lead to the cleavage of the peptide, leading to a release of fragments. Here, we used high-performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS) and pre-column online ortho-phthalaldehyde (OPA) derivatization-based amino acid analysis by HPLC with diode array detection and fluorescence detection to identify and quantify free amino acids released upon oxidation of proteins and peptides by hydroxyl radicals. Bovine serum albumin (BSA), ovalbumin (OVA) as model proteins, and synthetic tripeptides (comprised of varying compositions of the amino acids Gly, Ala, Ser, and Met) were used for reactions with hydroxyl radicals, which were generated by the Fenton reaction of iron ions and hydrogen peroxide. The molar yields of free glycine, aspartic acid, asparagine, and alanine per peptide or protein varied between 4 and 55%. For protein oxidation reactions, the molar yields of Gly (∼32-55% for BSA, ∼10-21% for OVA) were substantially higher than those for the other identified amino acids (∼5-12% for BSA, ∼4-6% for OVA). Upon oxidation of tripeptides with Gly in C-terminal, mid-chain, or N-terminal positions, Gly was preferentially released when it was located at the C-terminal site. Overall, we observe evidence for a site-selective formation of free amino acids in the OH radical-induced oxidation of peptides and proteins, which may be due to a reaction pathway involving nitrogen-centered radicals.
authors.affiliation: Multiphase Chemistry Department, Max Planck Institute for Chemistry, Hahn-Meitner-Weg 1, 55128, Mainz, Germany.
authors.forename: Fobang
authors.initials: F
authors.lastname: Liu
authors.affiliation: School of Environment and Energy, South China University of Technology, Higher Education Mega Center, Guangzhou, 510006, China.
authors.forename: Senchao
authors.initials: S
authors.lastname: Lai
authors.affiliation: Multiphase Chemistry Department, Max Planck Institute for Chemistry, Hahn-Meitner-Weg 1, 55128, Mainz, Germany.
authors.forename: Haijie
authors.initials: H
authors.lastname: Tong
authors.affiliation: Multiphase Chemistry Department, Max Planck Institute for Chemistry, Hahn-Meitner-Weg 1, 55128, Mainz, Germany.
authors.forename: Pascale S J
authors.initials: PSJ
authors.lastname: Lakey
authors.affiliation: Multiphase Chemistry Department, Max Planck Institute for Chemistry, Hahn-Meitner-Weg 1, 55128, Mainz, Germany.|Department of Chemistry, University of California, Irvine, 1102 Natural Sciences II, Irvine, CA, 92697-2025, USA.
authors.forename: Manabu
authors.initials: M
authors.lastname: Shiraiwa
authors.affiliation: Division 1.5 Protein Analysis, Federal Institute for Materials Research and Testing (BAM), Richard-Willstätter-Str. 11, 12489, Berlin, Germany.
authors.forename: Michael G
authors.initials: MG
authors.lastname: Weller
authors.affiliation: Multiphase Chemistry Department, Max Planck Institute for Chemistry, Hahn-Meitner-Weg 1, 55128, Mainz, Germany.
authors.forename: Ulrich
authors.initials: U
authors.lastname: Pöschl
authors.affiliation: Multiphase Chemistry Department, Max Planck Institute for Chemistry, Hahn-Meitner-Weg 1, 55128, Mainz, Germany. c.kampf@mpic.de.|Institute for Inorganic and Analytical Chemistry, Johannes Gutenberg University Mainz, Duesbergweg 10-14, 55128, Mainz, Germany. c.kampf@mpic.de.|Institute for Organic Chemistry, Johannes Gutenberg University Mainz, Duesbergweg 10-14, 55128, Mainz, Germany. c.kampf@mpic.de.
authors.forename: Christopher J
authors.initials: CJ
authors.lastname: Kampf
mesh_terms: D000596:Amino Acids / Q000737:chemistry*; D017665:Hydroxyl Radical / Q000737:chemistry*; D010084:Oxidation-Reduction; D010455:Peptides / Q000737:chemistry*; D011506:Proteins / Q000737:chemistry*; D017382:Reactive Oxygen Species / Q000737:chemistry
chemical_list: D000596:Amino Acids; D010455:Peptides; D011506:Proteins; D017382:Reactive Oxygen Species; D017665:Hydroxyl Radical
keywords: Amino acid analysis; HPLC-MS; Hydroxyl radicals; Oxidation; Peptides; Proteins