SUBSTRATE-SPECIFICITY AND INHIBITOR SENSITIVITY OF CA2+ S100 DEPENDENT TWITCHIN KINASES/

Myosin-associated giant protein kinases of the titin/twitchin-like sup erfamily have previously been implicated in the regulation of muscle f unction, based on genetic and physiological studies. We find that reco mbinant constitutively active Caenorhabditis elegans and Aplysia twitc hin kinase fragments differ in their catalytic activities and peptide- substrate specificities, as well as in their sensitivities to the naph thalene sulfonamide inhibitors oronaphthalenesulfonyl)-1H-hexahydro-1, 4-diazepine (ML-7) and odonaphthalenesulfonyl)-1H-hexahydro-1,4-diazep ine (ML-9). The constitutively active Aplysia twitchin kinase fragment has a remarkably high activity (V-max > 100 mu mol . min(-1) . mg(-1) ) towards some substrate peptides. The autoinhibited forms of these tw itchin kinases can be activated in a Ca2+-dependent manner by the dime ric form of the S100A1 protein (S100A1(2)). The twitchin kinase S100A1 (2)-binding site can also bind Ca2+/calmodulin but neither kinase is a ctivated by calmodulin. The data provide a functional basis for the on going crystallographic study of twitchin kinase fragments.