CHARACTERIZATION OF THE SECONDARY STRUCTURE OF CALMODULIN IN COMPLEX WITH A CALMODULIN-BINDING DOMAIN PEPTIDE

The interaction between calcium-saturated chicken calmodulin and a peptide corresponding to the calmodulin-binding domain of the chicken smooth muscle myosin light chain kinase has been studied by multinuclear and multidimensional nuclear magnetic resonance methods. Extensive H-1 and N-15 resonance assignments of calmodulin in the complex have been obtained from the analysis of two- and three-dimensional nuclear magnetic resonance spectra. The assignment of calmodulin in the complex was facilitated by the use of selective labeling of the protein with alpha-N-15-labeled valine, alanine, lysine, leucine, and glycine. These provided reference points during the main-chain-directed analysis of three-dimensional spectra of complexes prepared with uniformly N-15-labeled calmodulin. The pattern of nuclear Overhauser effects (NOE) seen among main-chain amide NH, C(alpha)H, and C(beta)H hydrogens indicates that the secondary structure of the globular domains of calmodulin in the complex closely corresponds to that observed in the calcium-saturated state of the protein in the absence of bound peptide. However, the backbone conformation of residues 76-84 adopts an "tended chain conformation upon binding of the peptide in contrast to its helical conformation in the absence of peptide. A sufficient number of NOEs between the globular domains of calmodulin and the bound peptide have been found to indicate that the N- and C-terminal regions of the peptide interact with the C- and N-terminal domains of calmodulin, respectively. The significance of these results are discussed in terms of recently proposed models for the structure of calmodulin-peptide complexes.