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Towards the identification of late‐embryogenic‐abundant phosphoproteome in Arabidopsis by 2‐DE and MS
Author(s) -
Irar Sami,
Oliveira Eliandre,
Pagès Montserrat,
Goday Adela
Publication year - 2006
Publication title -
proteomics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.26
H-Index - 167
eISSN - 1615-9861
pISSN - 1615-9853
DOI - 10.1002/pmic.200500387
Subject(s) - arabidopsis , thermostability , proteomics , chemistry , biochemistry , chromatography , mass spectrometry , phosphatase , peptide , storage protein , phosphorylation , arabidopsis thaliana , biology , enzyme , mutant , gene
Late‐embryogenesis‐abundant (LEA) proteins accumulate as plant seeds desiccate and also in vegetative organs during periods of stress. They are predicted to play a role in plant stress tolerance. In the present study, we have initiated the characterization of phosphorylated LEA proteins present in the Arabidopsis seed, using a strategy that combines the thermostability (solubility upon heating) of many LEA‐type proteins with the use of phosphoaffinity chromatography to obtain an enriched subpopulation of phosphoproteins. The specificity and efficiency of the procedure was assessed by alkaline phosphatase treatment and by a specific stain for phosphoproteins, in addition to the immunodetection of AtRab18, a phosphorylated LEA protein present in the mature dry seed. The phosphoproteins were identified by MS either by PMF using MALDI‐TOF MS after 2‐DE separation, or by peptide sequencing using both capillary LC MS/MS (LC μESI‐ITMS/MS) and nanoLC coupled to nanoESI‐MS/MS (LC‐nanoESI‐Q‐TOF‐MS/MS). Several LEA‐type and storage‐like proteins were identified as components of the phosphoproteome of the Arabidopsis seed.