Open AccessAllosteric Activation of Kinases: Design and Application of RapR Kinases
Author(s) -
Karginov Andrei V.,
Hahn Klaus M.
Publication year - 2011
Publication title -
current protocols in cell biology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.149
H-Index - 38
eISSN - 1934-2616
pISSN - 1934-2500
DOI - 10.1002/0471143030.cb1413s53
Subject(s) - kinase , allosteric regulation , sh3 domain , proto oncogene tyrosine protein kinase src , microbiology and biotechnology , protein kinase domain , protein serine threonine kinases , biology , biochemistry , chemistry , protein kinase a , enzyme , gene , mutant
Here we describe a method for the engineered regulation of protein kinases in living cells, the design and application of RapR (rapamycin regulated) kinases. The RapR kinase method enables activation of kinases with high specificity and precise temporal control. Insertion of an engineered allosteric switch, the iFKBP domain, at a structurally conserved position within the kinase catalytic domain makes the modified kinase inactive. Treatment with rapamycin or its non‐immunosuppressive analogs triggers interaction with a small FKBP‐rapamycin‐binding domain (FRB), restoring the activity of the kinase. The reagents used in this method are genetically encoded or membrane permeable, enabling ready application in many systems. Based on the structural similarity of kinase catalytic domains, this method is likely applicable to a wide variety of kinases. Successful regulation has already been demonstrated for three kinases representing both tyrosine and serine/threonine kinase families (p38, FAK, Src). Procedures for designing and testing RapR kinases are discussed. Curr. Protoc. Cell Biol . 53:14.13.1‐14.13.16. © 2011 by John Wiley & Sons, Inc.