DFG - PrKX

With 518 genes the protein kinases represent the second largest family of proteins in the human genome and have key responsibilities in the signal transduction and the regulation of metabolism in eukaryotic cells. Here, the cAMP-dependent protein kinase (PKA) is one of the best-characterized protein kinases. Despite the broad range of knowledge about the structure and biochemistry of PCA major isoforms (Cα, Cβ) the functionality of the human protein kinase catalytic isoform X (PrKX) has been described only in principle. PrKX has evolved phylogenetically to a separate PKA subfamily and shows an unusual regulatory mechanism.
Since there is no detailed information on the substrate specificity and specific interaction partners, (1) specific substrates should be marked and identifyed selectively via a chemical genetic approach and high resolution mass spectrometry. In addition, substrates shoul be identifyed by (2) peptide- and protein-arrays and binding partners shoul be identifyed (3) by a novel yeast two hybrid system and (4) a chemical proteomic approach respectively. The validation of potential binding partner take place in vitro via surface plasmon resonance (SPR) and in vivo via bioluminescence resonance energy transfer (BRET). Putative phosphorylation sites should be detected with highly specific antibody and verified by mass spectrometry.
These results should serve as a basis for further functional studies of the role of PrKX.

back