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

A1 Originalartikel i en vetenskaplig tidskrift (referentgranskad)


Interna författare/redaktörer


Publikationens författare: Kochin V, Imanishi SY, Eriksson JE
Publiceringsår: 2006
Tidskrift: Proteomics
Tidskriftsakronym: Proteomics
Volym: 6
Nummer: 21
Artikelns första sida, sidnummer: 5676
Artikelns sista sida, sidnummer: 5682
ISSN: 1615-9853


Abstrakt

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.

Senast uppdaterad 2020-26-02 vid 07:06