We isolated a constitutively active form of cGMP-dependent protein kinase Iα (cGK Iα) by PCR-driven random mutagenesis. The replacement of Ile-63 by Thr in the autoinhibitory domain results in the enhancement of autophosphorylation and the basal kinase activity in the absence of cGMP. The hydrophobicity at position 63 is essential for the inactive state of cGK Iα, and Ile-78 of cGK Iβ is also required for the autoinhibitory property. Furthermore, cGK Iα (Ile-63-Thr) is constitutively active in vivo. These findings suggest that a conserved residue in the autoinhibitory domain was involved in the autoinhibition of both cGK Is.
