A new potent calmodulin antagonist with arylalkylamine structure: Crystallographic, spectroscopic and functional studies

An arylalkylamine-type calmodulin antagonist, N-(3,3-diphenylpropyl)-N'-[1- R-(3,4-bis-butoxyphenyl)ethyl]-propylene-diamine (AAA) is presented and its complexes with calmodulin are characterized in solution and in the crystal . Near-UV circular dichroism spectra show that AAA binds to calmodulin with 2:1 stoichiometry in a Ca2+-dependent manner. The crystal structure with 2 :1 stoichiometry is determined to 2.64 Angstrom resolution. The binding of AAA causes domain closure of calmodulin similar to that obtained with trifl uoperazine. Solution and crystal data indicate that each of the two AAA mol ecules anchors in the hydrophobic pockets of calmodulin, overlapping with t wo trifluoperazine sites, i.e. at a hydrophobic pocket and an interdomain s ite. The two AAA molecules also interact with each other by hydrophobic for ces; A competition enzymatic assay has revealed that AAA inhibits calmoduli n-activated phosphodiesterase activity at two orders of magnitude lower con centration than trifluoperazine. The apparent dissociation constant of AAA to calmodulin is 18 nM, which is commensurable with that of target peptides . On the basis of the crystal structure, we propose that the high-affinity binding is mainly due to a favorable entropy term, as the AAA molecule make s multiple contacts in its complex with calmodulin. (C) 2000 Academic Press .