Identification of a specific amino acid cluster in the calmodulin-binding domain of the neuronal nitric oxide synthase.

The calmodulin (CaM) binding domain of rat neuronal nitric oxide synthase (nNOS) was analyzed using 3 synthetic peptides corresponding to different regions of the middle portion of the enzyme. One corresponding to nNOS 732-754 gave complete inhibition of NOS enzyme activity with an IC50 of about 1 microM. Kinetic analysis indicated that the inhibition was not competitive with respect to L-arginine and the peptide produced a Ca2+ dependent, electrophoretic mobility shift of CaM on 1 M urea gels. A specific hydrophobic/basic amino acid cluster in the rat nNOS sequence, Lys732 Lys Leu, that was critical for its CaM binding was also identified in this study.