Mutations in the catalytic subunit of the cAMP-dependent protein kinase interfere with holoenzyme formation without disrupting inhibition by protein kinase inhibitor.
Three amino acids were identified in the catalytic (C) subunit of the cyclic AMP-dependent protein kinase that are involved in interaction with regulatory (R) subunit, but not with the specific protein kinase inhibitor, PKI. In a functional assay for gene induction, a C expression vector with serine or arginine substituted for Leu-198 and the double mutant C, His-87-->Gln/Trp-196-->Arg (Orellana, S. A., and McKnight, G. S. (1992) Proc. Natl. Acad. Sci, U.S.A. 89, 4726-4730), retained activity in the presence of an excess of RI or RII. In contrast, cotransfection of a full-length PKI expression vector completely inhibited the activity of both mutant and wild type C subunits. These data suggest that although the substrate/pseudosubstrate sites of R and PKI interact with C at the catalytic site, there is an additional domain on the C subunit that is involved in holoenzyme formation with R subunit and is distinct from sites specifying high affinity PKI binding.
Orellana
S A
SA
Department of Pharmacology, University of Washington, Seattle 98195.
Amieux
P S
PS
Zhao
X
X
McKnight
G S
GS
D003198:Computer Simulation; D006801:Humans; D008958:Models, Molecular; D016296:Mutagenesis; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases / Q000235:genetics* / Q000378:metabolism; D013379:Substrate Specificity; D014407:Tumor Cells, Cultured
D047428:Protein Kinase Inhibitors; D011494:Protein Kinases