Identification of the calmodulin-binding domain of neuron-specific proteinkinase C substrate protein CAP-22/NAP-22 - Direct involvement of protein myristoylation in calmodulin-target protein interaction

Various proteins in the signal transduction pathways as well as those of vi ral origin have been shown to be myristoylated, Although the modification i s often essential for the proper functioning of the modified protein, the m echanism by which the modification exerts its effects is still largely unkn own. Brain-specific protein kinase C substrate, CAP-23/NAP-22, which is inv olved in the synaptogenesis and neuronal plasticity, binds calmodulin, but the protein lacks any canonical calmodulin-binding domain. In the present r eport, we show that CAP-23/NAP-22 isolated from rat brain is myristoylated and that the modification is directly involved in its interaction with calm odulin. Myristoylated and non-myristoylated recombinant proteins were produ ced in Escherichia coli, and their calmodulin-binding properties were exami ned. Only the former bound to calmodulin, Synthetic peptides based on the N -terminal sequence showed similar binding properties to calmodulin, only wh en they were myristoylated, The calmodulin-binding site narrowed down to th e myristoyl moiety together with a nine-amino acid N-terminal basic domain. Phosphorylation of a single serine residue in the N-terminal domain (Ser(5 )) by protein kinase C abolished the binding. Furthermore, phosphorylation of CAP-23/NAP-22 by protein kinase C was also found myristoylation-dependen t, suggesting the importance of myristoylation in protein-protein interacti ons.