Genetic selection of regulatory mutants of mammalian adenylyl cyclases

This chapter describes a genetic system utilizing the expression of mammalian type V adenylyl cyclase in yeast and its use in isolating mutant adenylyl cyclases defective in their regulation by G s α. X-ray crystallography has opened an exciting window into the physical chemistry of adenylyl cyclase. The three-dimensional structures that have been determined of individual domains and complexes of the essential catalytic core with activators and substrate analogs have provided important insights into the mechanism of catalysis and activation. Described here are the procedures used to generate crystals of the adenylyl cyclase catalytic core bound to its activators Gα (stimulatory G protein α subunit) and forskolin.