Resolution of adenylate cyclase sensitive and insensitive to Ca²⁺ and calcium-dependent regulatory protein (CDR) by CDR-Sepharose affinity chromatography

Partially purified adenylate cyclase [ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1] from bovine brain cortex was fractionated into two separate forms by calcium-dependent regulatory protein (CDR)-Sepharose affinity chromatography. The major form of the enzyme, comprising ~80% of the applied activity, did not bind to the affinity column in the presence of Ca²⁺ and was insensitive to the CDR. Approximately 20% of adenylate cyclase activity was absorbed to CDR-Sepharose in the presence of Ca²⁺. This activity was stimulated by Ca²⁺ and CDR. This study directly demonstrates that brain cortex contains Ca²⁺.CDR-sensitive and -insensitive forms of adenylate cyclase and indicates that CDR-Sepharose may be a useful tool for purification of adenylate cyclase. The Ca²⁺-stimulated adenylate cyclase was purified at least 55-fold with a 13% yield.