title: New protein isoforms identified within Arabidopsis thaliana seed oil bodies combining chymotrypsin/trypsin digestion and peptide fragmentation analysis.
abstract: Plant seed oil bodies, subcellular lipoprotein inclusions providing storage reserves, are composed of a neutral lipid core surrounded by a phospholipid monolayer with several integrated proteins that play a significant role in stabilization of the particles and probably also in lipid mobilization. Oil bodies' proteins are generally very hydrophobic, due to the long uncharged sequences anchoring them into the lipid core, which makes them extremely difficult to handle and to digest successfully. Although oil bodies have been intensively studied during last decades, not all their proteins have been identified yet. To overcome the problems connected with their identification, a method based on SDS-PAGE, in-gel digestion and LC-MS/MS analysis was used. Digestion was carried out with trypsin and chymotrypsin, single or in combination, which increased significantly the number of identified peptides, namely the hydrophobic ones. Thanks to this methodology it was possible to achieve an extensive coverage of proteins studied, to analyze their N-terminal modifications and moreover, to detect four new oil bodies' protein isoforms, which demonstrates the complexity of oil bodies' protein composition.
authors.affiliation: Institute of Chemical Technology Prague, Department of Biochemistry and Microbiology, Prague, Czech Republic. martina.vermachova@vscht.cz
authors.forename: Martina
authors.initials: M
authors.lastname: Vermachova
authors.forename: Zita
authors.initials: Z
authors.lastname: Purkrtova
authors.forename: Jiri
authors.initials: J
authors.lastname: Santrucek
authors.forename: Pascale
authors.initials: P
authors.lastname: Jolivet
authors.forename: Thierry
authors.initials: T
authors.lastname: Chardot
authors.forename: Milan
authors.initials: M
authors.lastname: Kodicek
mesh_terms: D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000737:chemistry* / Q000378:metabolism; D029681:Arabidopsis Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D002853:Chromatography, Liquid; D002918:Chymotrypsin / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000032:analysis* / Q000737:chemistry; D010938:Plant Oils / Q000737:chemistry; D020033:Protein Isoforms / Q000737:chemistry* / Q000145:classification / Q000302:isolation & purification; D012639:Seeds / Q000737:chemistry* / Q000378:metabolism; D020539:Sequence Analysis, Protein; D014357:Trypsin / Q000737:chemistry; D014617:Vacuoles / Q000737:chemistry*
chemical_list: D029681:Arabidopsis Proteins; D010446:Peptide Fragments; D010938:Plant Oils; D020033:Protein Isoforms; D002918:Chymotrypsin; D014357:Trypsin