INTERACTION OF THE RECOMBINANT S100A1 PROTEIN WITH TWITCHIN KINASE, AND COMPARISON WITH OTHER CA2-BINDING PROTEINS()

The giant myosin-associated twitchin kinase, a member of the Ca2+-regu lated protein kinase superfamily, is activated by the EF-hand protein S100A1 in a Ca2+-dependent and Zn2+-enhanced manner. We used recombina nt S100A1 to further characterize the interaction between the two prot eins. Zn2+ enhanced the binding of Ca2+/S100A1 to twitchin kinase frag ments (K-d < 50 nM) in assays using a BIAcore biosensor by reducing th e S100A1 off rate. Other Ca2+-binding proteins (S100A6, calmodulin, an d the calmodulin-like domain of Ca2+-dependent protein kinase abound t o the kinase but did not activate it. These results indicate that bind ing of Ca2+-binding proteins alone is insufficient to trigger the intr amolecular rearrangement of kinase autoinhibitory contacts required fo r twitchin kinase activation that is specifically elicited by the S100 A1 protein. Kinase fragments that contained only the autoinhibited cat alytic sequence or an additional immunoglobulin-like domain had very s imilar properties, indicating that the tethered immunoglobulin-like do main does not modulate kinase regulation.