Enrichment of phosphoproteins and phosphopeptide derivatization identify universal stress proteins in elicitor-treated Arabidopsis.
Protein phosphorylation is a key biological process that regulates reactions involved in plant-microbe interactions. The phosphorylated form of a protein often represents only a small fraction of the total population and can be problematic to analyze in a mass spectrometer. We demonstrate how a titanium dioxide (TiO(2)) resin can be employed for the enrichment of phosphoproteins, as well as a method to derivatize TiO(2)-purified phosphopeptides to facilitate determination of the exact site of phosphorylation. The use of these methods was exemplified by the identification of two plant proteins that were shown to be phosphorylated after the elicitation of Arabidopsis cells with Phytophthora infestans zoospores and xylanase. Both of the proteins that were identified, At5g54430.1 and At4g27320.1, were found to contain a universal stress protein domain with conserved residues for ATP binding.
Lenman
Marit
M
Department of Cell and Organism Biology, Lund University, Lund, Sweden.
Sörensson
Carolin
C
Andreasson
Erik
E
D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000166:cytology / Q000378:metabolism* / Q000382:microbiology; D029681:Arabidopsis Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D002853:Chromatography, Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D018506:Gene Expression Regulation, Plant / Q000187:drug effects; D054884:Host-Pathogen Interactions; D008969:Molecular Sequence Data; D010748:Phosphopeptides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010750:Phosphoproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D010838:Phytophthora / Q000502:physiology; D017386:Sequence Homology, Amino Acid; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014025:Titanium / Q000737:chemistry / Q000494:pharmacology
D029681:Arabidopsis Proteins; D010748:Phosphopeptides; D010750:Phosphoproteins; C009495:titanium dioxide; D014025:Titanium