Production of recombinant plant calmodulin and its use to detect calmodulin-binding proteins

Calmodulin (CaM) is a major intracellular receptor for second messenger Ca2+ signals and is expressed in every cell of all eukaryotes, including plants. CaM transduces these signals by binding four Ca2+, and binding to and altering the activities of a variety of enzymatic, cytoskeletal, and structural proteins. This chapter presents various methods to detect plant CaM-binding proteins that exploit the simplicity with which recombinant plant CaM can be purified in milligram amounts from modest-sized cultures of induced Escherichia coli and labeled with a variety of reporters, which offer the advantages of sensitive detection of CaM-binding proteins and relatively low toxicity. The utility of these methods described for labeling CaM to use as a probe for detecting CaM-binding proteins can be illustrated by using two applications, which involve detecting CaM-binding activity in protein fractions after separation by sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS–PAGE) and in induced lysates of recombinant bacteriophage harboring complementary DNA (cDNA) expression libraries. © 1995, Academic Press Inc.