title: Characterization of antimicrobial histone sequences and posttranslational modifications by mass spectrometry.
abstract: Histones typically play a role in DNA packaging and transcription regulation. These proteins are heavily modified by acetylation, methylation, phosphorylation and/or ubiquitination, and various combinations of these modifications alter histone functions and form the basis of the histone code. Furthermore, histones, including those found in shrimp, have recently been found to possess antimicrobial properties; however, the sequences and posttranslational modifications of shrimp histones are largely unknown. In this study mass spectrometry was used to characterize the primary structure of the shrimp antimicrobial histone. A combination of in-solution digestion and in-gel propionylation/digestion followed by LC-MS-MS and MALDI-TOF-TOF analysis was used. Over 80% of each histone sequence was obtained by in-solution digestion; however, none of the N-terminal domains was sequenced with this method. An in-gel propionylation method was optimized to recover and sequence the extremely hydrophilic histone N-termini. This method was then applied to shrimp hemocyte lysates separated on a 1-D SDS-PAGE gel. Overall, 95% coverage was obtained for the histone sequences as well as the identification of posttranslational sites such as acetylation, methylation and phosphorylation.
authors.affiliation: Medical University of South Carolina, Department of Cell and Molecular Pharmacology, Charleston, SC 29425, USA.
authors.forename: Séverine A
authors.initials: SA
authors.lastname: Ouvry-Patat
authors.forename: Kevin L
authors.initials: KL
authors.lastname: Schey
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000890:Anti-Infective Agents / Q000737:chemistry*; D004591:Electrophoresis, Polyacrylamide Gel; D006434:Hemocytes / Q000737:chemistry*; D006657:Histones / Q000737:chemistry*; D008969:Molecular Sequence Data; D033561:Penaeidae / Q000737:chemistry*; D011499:Protein Processing, Post-Translational; D020539:Sequence Analysis, Protein; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D000890:Anti-Infective Agents; D006657:Histones