title: A versatile method for deciphering plant membrane proteomes.
abstract: Proteomics is a very powerful approach to link the information contained in sequenced genomes, like that of Arabidopsis, to the functional knowledge provided by studies of plant cell compartments. This article summarizes the different steps of a versatile strategy that has been developed to decipher plant membrane proteomes. Initiated with envelope membranes from spinach chloroplasts, this strategy has been adapted to thylakoids, and further extended to a series of membranes from the model plant Arabidopsis: chloroplast envelope membranes, plasma membrane, and mitochondrial membranes. The first step is the preparation of highly purified membrane fractions from plant tissues. The second step in the strategy is the fractionation of membrane proteins on the basis of their physico-chemical properties. Chloroform/methanol extraction and washing of membranes with NaOH, NaCl or any other agent led to the simplification of the protein content of the fraction to be analysed. The next step is the genuine proteomic step, i.e. the separation of proteins by 1D-gel electrophoresis followed by in-gel proteolytic digestion of the polypeptides, analysis of the proteolytic peptides using mass spectrometry, and protein identification by searching through databases. The last step is the validation of the procedure by checking the subcellular location. The results obtained by using this strategy demonstrate that a combination of different proteomics approaches, together with bioinformatics, indeed provide a better understanding of the biochemical machinery of the different plant membranes at the molecular level.
authors.affiliation: Laboratoire de Physiologie Cellulaire Végétale, UMR 5168 CEA/CNRS/INRA/Université Joseph Fourier, CEA-Grenoble, F-38054 Grenoble-cedex 9, France.
authors.forename: Norbert
authors.initials: N
authors.lastname: Rolland
authors.forename: Myriam
authors.initials: M
authors.lastname: Ferro
authors.forename: Geneviève
authors.initials: G
authors.lastname: Ephritikhine
authors.forename: Anne
authors.initials: A
authors.lastname: Marmagne
authors.forename: Claire
authors.initials: C
authors.lastname: Ramus
authors.forename: Sabine
authors.initials: S
authors.lastname: Brugière
authors.forename: Daniel
authors.initials: D
authors.lastname: Salvi
authors.forename: Daphné
authors.initials: D
authors.lastname: Seigneurin-Berny
authors.forename: Jacques
authors.initials: J
authors.lastname: Bourguignon
authors.forename: Hélène
authors.initials: H
authors.lastname: Barbier-Brygoo
authors.forename: Jacques
authors.initials: J
authors.lastname: Joyard
authors.forename: Jérome
authors.initials: J
authors.lastname: Garin
mesh_terms: D000468:Alkalies / Q000494:pharmacology; D017360:Arabidopsis / Q000378:metabolism / Q000648:ultrastructure*; D029681:Arabidopsis Proteins / Q000302:isolation & purification* / Q000378:metabolism; D002458:Cell Fractionation; D019295:Computational Biology; D007425:Intracellular Membranes / Q000378:metabolism*; D008565:Membrane Proteins / Q000302:isolation & purification* / Q000378:metabolism; D040901:Proteomics / Q000379:methods*; D012492:Salts / Q000494:pharmacology
chemical_list: D000468:Alkalies; D029681:Arabidopsis Proteins; D008565:Membrane Proteins; D012492:Salts