Activating Mutation of Adenylyl Cyclase Reverses its Inhibition by G Proteins

We have implemented a yeast genetic selection developed previously by our laboratory to identify mutant mammalian type V adenylyl cyclases insensitive to inhibition by Gia. One muta-tion isolated was localized to the first cytoplasmic domain at a Phe residue (position 400), which is conserved in all nine iso-forms of membrane-bound mammalian adenylyl cyclase. Bio-chemical characterization of the F400Y mutant revealed a dra-matic conversion of the Gia response from inhibitory to stimulatory. This mutation results in additional activating ef-fects. The mutant exhibits an enhanced sensitivity toward ac-tivation by either Gsa or forskolin. Synergism between Gsa and forskolin is not observed for the F400Y mutant, presumably because the mutant already is in the sensitized state. Addition-ally, an enhancement of the basal unstimulated activity wa