| Understanding plant mechanisms under nutrient limitation or pollution are of great interest for agriculture and phytoremediation purposes. The cDNA sequence for a novel A&barbelow;rabidopsis t&barbelow;haliana Ser/Thr protein kinase, belonging to the p&barbelow;hosphate-s&barbelow;tarvation r&barbelow;esponsive gene group 1&barbelow; (Atpsr1), has been cloned into the pGEX-His Escherichia coli expression vector. AtPSR1 belongs to the SNF1-related protein kinase 2 (SnRK2) subfamily and contains a unique acidic C-terminus of unknown function. Northern blots have indicated that Atpsr1 is transcribed in root tissue and that transcription is increased under Pi starvation. GST-AtPSR1, expressed in E. coli, was affinity purified in the presence of sarkosyl. A far-western overlay assay, probed with GST-AtPSR1, identified m&barbelow;yosin h&barbelow;eavy c&barbelow;hain (MHC) as a potential substrate. Phosphoamino acid analysis, by two-dimensional gel electrophoresis, demonstrated that phosphorylation occurred on serine residues. The activity of GST-AtPSR1 was analyzed by quantification of the phosphorylated 25 kDa protein. A series of mutations were analyzed for their effects on kinase activity. (Abstract shortened by UMI.)... |
