title: The membrane proteome of Halobacterium salinarum.
abstract: The identification of 114 integral membrane proteins from Halobacterium salinarum was achieved using liquid chromatography/tandem mass spectrometric (LC/MS/MS) techniques, representing 20% of the predicted alpha-helical transmembrane proteins of the genome. For this experiment, a membrane preparation with only minor contamination by soluble proteins was prepared. From this membrane preparation a number of peripheral membrane proteins were identified by the classical two dimensional gel electrophoresis (2-DE) approach, but identification of integral membrane proteins largely failed with only a very few being identified. By use of a fluorescently labeled membrane preparation, we document that this is caused by an irreversible precipitation of the membrane proteins upon isoelectric focusing (IEF). Attempts to overcome this problem by using alternative IEF methods and IEF strip solubilisation techniques were not successful, and we conclude that the classical 2-DE approach is not suited for the identification of integral membrane proteins. Computational analysis showed that the identification of integral membrane proteins is further complicated by the generation of tryptic peptides, which are unfavorable for matrix assisted laser desorption/ionization time of flight mass spectrometric peptide mass fingerprint analysis. Together with the result from the analysis of the cytosolic proteome (see preceding paper), we could identify 34% (943) of all gene products in H. salinarum which can be theoretically expressed. This is a cautious estimate as very stringent criteria were applied for identification. These results are available under www.halolex.mpg.de.
authors.affiliation: Abteilung für Membranbiochemie, Max-Planck-Institut für Biochemie, D-82152 Martinsried, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Klein
authors.forename: Carolina
authors.initials: C
authors.lastname: Garcia-Rizo
authors.forename: Birgit
authors.initials: B
authors.lastname: Bisle
authors.forename: Beatrix
authors.initials: B
authors.lastname: Scheffer
authors.forename: Hans
authors.initials: H
authors.lastname: Zischka
authors.forename: Friedhelm
authors.initials: F
authors.lastname: Pfeiffer
authors.forename: Frank
authors.initials: F
authors.lastname: Siedler
authors.forename: Dieter
authors.initials: D
authors.lastname: Oesterhelt
mesh_terms: D002462:Cell Membrane / Q000737:chemistry*; D002499:Centrifugation, Density Gradient; D002853:Chromatography, Liquid; D015180:Electrophoresis, Gel, Two-Dimensional; D004591:Electrophoresis, Polyacrylamide Gel; D019617:Halobacterium salinarum / Q000737:chemistry*; D007525:Isoelectric Focusing; D013058:Mass Spectrometry; D008565:Membrane Proteins / Q000737:chemistry* / Q000302:isolation & purification; D009994:Osmolar Concentration; D020543:Proteome / Q000032:analysis*
chemical_list: D008565:Membrane Proteins; D020543:Proteome