Fast track to a phosphoprotein sketch - MALDI-TOF characterization of TLC-based tryptic phosphopeptide maps at femtomolar detection sensitivity

A1 Journal article (refereed)


Internal Authors/Editors


Publication Details

List of Authors: Kochin V, Imanishi SY, Eriksson JE
Publication year: 2006
Journal: Proteomics
Journal acronym: Proteomics
Volume number: 6
Issue number: 21
Start page: 5676
End page: 5682
ISSN: 1615-9853


Abstract

Tryptic phosphopeptide mapping by TLC on microcrystalline cellulose has been a convenient method to get a fast and highly reproducible overview of the number of phosphopeptides present in any given (32)P-labeled phosphoprotein. This method also provides an immediate presentation of the relative phosphorylation stoichiometry between individual phosphopeptides. However, so far, traditional tryptic phosphopeptide maps have not been useful for phosphoproteomics applications, as the S/N has been very poor, due to the large number of quenching substances and contaminants present on cellulose plates. In this study, we present a rapid and easy method for phosphopeptides identification from 2-D phosphopeptide maps (2-D-PPMs). We obtain improved sensitivity (femtomole levels) upon MALDI-TOF MS analysis of phosphopeptides extracted from 2-D-PPMs. Using this approach we could confidently characterize the major phosphorylation sites of in vivo and in vitro (32)P-labeled proteins.

Last updated on 2019-21-10 at 02:56