A synthetic analog of the calmodulin-binding domain of myosin light chain kinase inhibits melanotropin receptor function and activation of adenylate cyclase.

In this study a synthetic analog of the calmodulin-binding domain of myosin light chain kinase, a 17-amino-acid peptide (M5) was used to examine the possible role of calmodulin in melanotropin receptor function. Binding of beta-melanocyte-stimulating hormone to its membrane receptor and subsequent stimulation of adenylate cyclase (AC) were found to be specifically inhibited by M5 in a dose-dependent and noncompetitive manner, both in intact M2R melanoma cells and in a plasma membrane preparation derived thereof. Half-maximal inhibition of both hormone binding and melanotropin-sensitive AC activity was shown to occur at approximately 1 microM M5. In contrast, stimulation of AC by prostaglandin E1, guanosine 5'-O-(3-thio)triphosphate, forskolin, and unstimulated enzyme activity were unaffected by the presence of M5, under the same assay conditions. Furthermore, addition of a molar excess of calmodulin to the AC assay completely abolished the inhibitory effects of the peptide on melanotropin-stimulated AC activity. Other peptides of similar size, which bind to calmodulin with low affinity (e.g. glucagon, somatostatin, and vasoactive intestinal peptide), were shown to be totally ineffective in inhibiting melanotropin-sensitive AC. These findings, along with those shown previously for other antagonists of calmodulin, suggest a role for an M5-binding protein, as of yet unidentified, involved in the regulation of the melanotropin receptor function.