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Linking protein fractionation with multidimensional monolithic reversed-phase peptide chromatography/mass spectrometry enhances protein identification from complex mixtures even in the presence of abundant proteins.
Wienkoop, Stefanie; Glinski, Mirko; Tanaka, Nobuo; Tolstikov, Vladimir; Fiehn, Oliver; Weckwerth, Wolfram.
Afiliación
  • Wienkoop S; Max-Planck-Institut für Molekulare Pflanzenphysiologie, 14424 Potsdam, Germany.
Rapid Commun Mass Spectrom ; 18(6): 643-50, 2004.
Article en En | MEDLINE | ID: mdl-15052571
ABSTRACT
Recently, multidimensional shotgun proteomics has proven to be an alternative technology able to identify hundreds of proteins from single samples. Two major limitations of the technology are the presence of high abundance proteins (e.g. RUBISCO in plant leaf tissue) and the enormous number of co-eluting peptides that overstrain the loading and resolving capacity of conventional particle-packed columns as well as the capacity of electrospray ionisation due to ion suppression. Here, the coupling of fast performance liquid chromatography (FPLC) pre-fractionation of an Arabidopsis leaf protein extract and subsequent two-dimensional liquid chromatography/mass spectrometry with improved resolution using a monolithic silica C18 capillary column allowed the identification of 1032 unique proteins in a single 4 mg total protein plant leaf tissue sample. The reassignment of peptide IDs to distinct FPLC protein fractions enhances the identification procedure, especially in the case of present protein isoforms. The proposed strategy is useful to detect proteins otherwise not seen in conventional multidimensional chromatography/mass spectrometry approaches.
Asunto(s)
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Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Proteínas de Plantas / Cromatografía Líquida de Alta Presión / Cromatografía por Intercambio Iónico / Proteoma / Espectrometría de Masa por Ionización de Electrospray Tipo de estudio: Diagnostic_studies Idioma: En Revista: Rapid Commun Mass Spectrom Año: 2004 Tipo del documento: Article País de afiliación: Alemania
Buscar en Google
Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Proteínas de Plantas / Cromatografía Líquida de Alta Presión / Cromatografía por Intercambio Iónico / Proteoma / Espectrometría de Masa por Ionización de Electrospray Tipo de estudio: Diagnostic_studies Idioma: En Revista: Rapid Commun Mass Spectrom Año: 2004 Tipo del documento: Article País de afiliación: Alemania