We describe here mutations of the catalytic subunit alpha of protein kinase A (PKA) that introduce amino acid side chains into the ATP binding site and progressively transform the pocket to mimic that of Aurora protein kinases. The resultant PKA variants are enzymatically active and exhibit high affinity for ATP site inhibitors that are specific for Aurora kinases. These features make the Aurora-chimeric PKA a valuable tool for structure based drug discovery tasks. Analysis of crystal structures of the chimera reveal the roles of individual amino acid residues in the binding of a variety of inhibitors, offering key insights into selectivity mechanisms. Furthermore the high affinity for Aurora kinase specific inhibitors, combined with the favorable crystallizability properties of PKA allow rapid determination of inhibitor complex structures at atomic resolution. We demonstrate the utility of the Aurora-chimeric PKA by measuring binding kinetics for three Aurora kinase specific inhibitors, and present the X ray structures of the chimeric enzyme in complex with VX 680 (MK 0457) and JNJ 7706621 (Aurora kinase/Cdk inhibitor).
Mutants of protein kinase A that mimic the ATP-binding site of Aurora kinase.,Pflug A, de Oliveira TM, Bossemeyer D, Engh RA Biochem J. 2011 Jul 21. PMID:21774789
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↑ Pflug A, de Oliveira TM, Bossemeyer D, Engh RA. Mutants of protein kinase A that mimic the ATP-binding site of Aurora kinase. Biochem J. 2011 Jul 21. PMID:21774789 doi:10.1042/BJ20110592