Kinase probes

MolokanovaE, Kramer RH, Mechanism of inhibition of cyclic nucleotide-gated channel by protein tyrosine kinase probed with genistein, J Gen Physiol 117 219-234, 2001. 

V.T. Skamnaki, D.J. Owen, M.E. Noble, E.D. Lowe, G. Lowe, N.G. Oikonomakos, and L.N. Johnson. 1999. Catalytic mechanism of phosphorylase kinase probed by mutational studies Biochemistry 38 14718-14730. (PubMed)... 

Starting Points for Kinase Probe Discovery The benefits of chemical connectivity... 

Sox-based kinase probes report activity in continuous assays, are compatibie with ceii iysates and tissue homogenates, and can be used for smaii-moiecuie inhibitor screening. Upon design and synthesis, kinetic parameters for each Sox-peptide are determined and kinase seiectivity is addressed. Subsequentiy, sensors are evaiuated in ceii-iysate-based systems in the presence and absence of seiective inhibitors to estabiish seiectivity in compiex target sampies. 

Group II assays consist of those monitoring cellular second messengers. Thus, activation of receptors to cause Gs-protein activation of adenylate cyclase will lead to elevation of cytosolic or extracellularly secreted cyclic AMP. This second messenger phosphorylates numerous cyclic AMP-dependent protein kinases, which go on to phosphorylate metabolic enzymes and transport and regulatory proteins (see Chapter 2). Cyclic AMP can be detected either radiometrically or with fluorescent probe technology. 

Wang S, Wang X, Shi W et al (2008) Detection of local polarity and conformational changes at the active site of rabbit muscle creatine kinase with a new arginine-specific fluorescent probe. Biochim Biophys Acta 1784 415 -22... 

As in all eukaryotic cells, protein kinases play an important role in the life cycle of the kinetoplastids and, as such, are attractive targets. Recent efforts, predominantly through genetic (RNAi) means, have validated a number of kinases as essential for survival of T. brucei, but few have been explored with chemical probes [33-35]. 

Since kinases are not hydrolytic enzymes, a small molecule-based FRET probe does not seem to be a straight forward solution for this enzyme activity. Nevertheless, quite a number of fluorescent probes based on small substrate peptides have been prepared in... 

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