ARGININE-75 IN THE PSEUDOSUBSTRATE SEQUENCE OF TYPE I-BETA CGMP-DEPENDENT PROTEIN-KINASE IS CRITICAL FOR AUTOINHIBITION, ALTHOUGH AUTOPHOSPHORYLATED SERINE-63 IS OUTSIDE THIS SEQUENCE

Autoinhibitory domains in many protein kinases include either a phosph orylatable substrate-like sequence or a pseudosubstrate sequence, This study shows that I beta cGMP-dependent protein kinase (cGK) autophosp horylates Ser-63, which is in an atypical cGK substrate sequence (-(59 )AQKQSAS-) that is amino-terminal to the pseudosubstrate motif (-(74)K RQAI-), cGMP increases the rate of autophosphorylation (similar to 0,8 phosphate/cGK monomer) similar to 3-fold, Autophosphorylation is an i ntramolecular process since it is independent of cGK concentration, cG MP activation of cGK enhances proteolysis within and near the pseudosu bstrate site; treatment of dimeric cGK with three proteases produces t hree cGK monomers (similar to 67-70 kDa each), Their amino-terminal se quences are (75)RQAISAEPT-, (76)QAlSAEPTAF-, and (86)DIQDLSXV-, respec tively, cGMP stimulates these kinases by 10-, 2.5-, and 1,4-fold, resp ectively, compared with a 10-fold effect on intact cGK, Increased basa I activity accounts for the diminished stimulation, Thus, the primary autophosphorylation site of I beta cGK is well outside the pseudosubst rate site, but Arg-75 in the pseudosubstrate site is critical for auto inhibition, Autoinhibition also involves elements that are carboxyl-te rminal to Arg-75.