Planta

, Volume 229, Issue 5, pp 1123–1134

Phosphorylation site mapping of soluble proteins: bioinformatical filtering reveals potential plastidic phosphoproteins in Arabidopsis thaliana

  • Katharina Lohrig
  • Bernd Müller
  • Joulia Davydova
  • Dario Leister
  • Dirk Andreas Wolters
Original Article

DOI: 10.1007/s00425-009-0901-y

Cite this article as:
Lohrig, K., Müller, B., Davydova, J. et al. Planta (2009) 229: 1123. doi:10.1007/s00425-009-0901-y

Abstract

Protein phosphorylation is a major mode of regulation of metabolism, gene expression, and cell architecture. A combination of phosphopeptide enrichment strategies based on TiO2 and IMAC in addition to our MudPIT strategy revealed the detection of 181 phosphorylation sites which are located on 125 potentially plastidic proteins predicted by GoMiner, TargetP/Predotar in Arabidopsis thaliana. In our study phosphorylation on serine is favored over threonine and this in turn over phosphorylation on tyrosine residues, showing a percentage of 67.4% to 24.3% to 8.3% for pS:pT:pY. Four phosphorylated residues (S208, Y239, T246 and T330), identified by our approach have been fitted to the structure of the activated form of spinach RuBisCO, which are located in close proximity to the substrate binding site for ribulosebisphosphate. Potentially, these phosphorylation sites exert a direct influence on the catalytic activity of the enzyme. Such examples show nicely the value of the presented mass spectrometric dataset for further biochemical applications, since alternative mutation analysis often turns out to be unsuccessful, caused by mutations in essential proteins which result in lethal phenotypes.

Keywords

Arabidopsis Chloroplast Immobilized metal ion affinity chromatography (IMAC) TiO2 Multidimensional protein identification technology (MudPIT) Phosphorylation 

Abbreviations

IMAC

Immobilized metal ion affinity chromatography

MudPIT

Multidimensional protein identification technology

RP

Reversed phase

RuBisCo

Ribulose-1,5-bisphosphate carboxylase

SCX

Strong cation exchange

Supplementary material

425_2009_901_MOESM1_ESM.xls (179 kb)
Supplementary Table 1 (XLS 179 kb)

Copyright information

© Springer-Verlag 2009

Authors and Affiliations

  • Katharina Lohrig
    • 1
  • Bernd Müller
    • 2
  • Joulia Davydova
    • 2
  • Dario Leister
    • 3
  • Dirk Andreas Wolters
    • 1
    • 4
  1. 1.Department of Analytical ChemistryRuhr-University BochumBochumGermany
  2. 2.Mass Spectrometry Unit, Department Biology ILudwig-Maximilians-Universität MünchenMunichGermany
  3. 3.Lehrstuhl für Botanik, Department Biology ILudwig-Maximilians-Universität MünchenMunichGermany
  4. 4.Ruhr-University BochumBochumGermany