Preparation of Fluorescent, Cross-Linking, and Biotinylated Calmodulin Derivatives and Their Use in Studies of Calmodulin-Activated Phosphodiesterase and Protein Phosphatase

This chapter summarizes work that have carried out with three derivatives of calmodulin (CaM), each of which has unique advantages for the investigation of structure and function: 5-dimethylaminonaphthalene-1-sulfonyl(dansyl)-CaM, a fluorescent derivative; 3-(2-pyridyldithio)propionyl(PDP)-CaM, an activated sulfhydryl cross-linking derivative; and biotinyl-ε-aminocaproyl(Bio)-CaM, a biotin-containing derivative that forms stable complexes with avidin-enzyme conjugates. As the primary amino acid sequence of CaM is known, the site(s) of modification have, in some instances, been exactly defined using proteolytic and chemical cleavage of the modified protein. Thus, chemical modification can provide a valuable adjunct to classical physical methods for investigation of CaM structure and function. In addition to methods for their syntheses, procedures for characterization and use are outlined in the chapter. Properties and uses of Bio-CaM are also summarized in the chapter.