title: Proteolytic 18O labeling by peptidyl-Lys metalloendopeptidase for comparative proteomics.
abstract: The potential capabilities of a new proteolytic 18O labeling method employing peptidyl-Lys metalloendopeptidase (Lys-N) have been demonstrated for use in comparative proteomics. Conditions (pH>or=9.5) have been found such that Lys-N incorporates only a single 18O atom into the carboxyl terminus of each proteolytically generated peptide. This 18O labeling method has a major advantage over current protelytic 18O labeling methods that generate a mixture of isotopic isoforms resulting from the incorporation of one or two 18O atoms into each peptide species by the proteases (trypsin, Lys-C, or Glu-C) used. We demonstrate that the single 18O atom incorporation property of Lys-N overcomes the major problem of the current proteolytic 18O labeling methods and provides accurate quantification results for isotopically labeled peptides.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of North Dakota School of Medicine, Grand Forks, North Dakota 58203, USA.
authors.forename: K C Sekhar
authors.initials: KC
authors.lastname: Rao
authors.forename: Ryan T
authors.initials: RT
authors.lastname: Carruth
authors.forename: Masaru
authors.initials: M
authors.lastname: Miyagi
mesh_terms: D000595:Amino Acid Sequence; D002853:Chromatography, Liquid; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D013058:Mass Spectrometry; D008666:Metalloendopeptidases / Q000737:chemistry*; D008969:Molecular Sequence Data; D010103:Oxygen Isotopes / Q000737:chemistry*; D040901:Proteomics
chemical_list: D010103:Oxygen Isotopes; D008666:Metalloendopeptidases; C039830:peptidyl-Lys metalloendopeptidase