IDENTIFICATION OF COMMON AND DISTINCT RESIDUES INVOLVED IN THE INTERACTION OF ALPHA(I2) AND ALPHA(S) WITH ADENYLYL-CYCLASE

The G protein alpha subunits, alpha(s) and alpha(i2), have stimulatory and inhibitory effects, respectively, on a common effector protein, a denylyl cyclase, These effects require a GTP-dependent conformational change that involves three alpha subunit regions (Switches I-III). alp ha(s) residues in three adjacent loops, including Switch II, specify a ctivation of adenylyl cyclase. The adenylyl cyclase-specifying region of alpha(i2) is located within a 78-residue segment that includes two of these loops but none of the conformational switch regions, We have used an alanine-scanning mutagenesis approach within Switches I-III an d the 78-residue segment of alpha(i2) to identify residues required fo r inhibition of adenylyl cyclase, We found a cluster of conserved resi dues in Switch II in which substitutions cause major losses in the abi lities of both alpha(i2) and alpha(s) to modulate adenylyl cyclase act ivity but do not affect alpha subunit expression or the GTP-induced co nformational change, We also found two regions within the 78-residue s egment of alpha(i2) in which substitutions reduce the ability of alpha (i2) to inhibit adenylyl cyclase, one of which corresponds to an effec tor-activating region of alpha(s). Thus, both alpha(i2) and alpha(s) i nteract with adenylyl cyclase using: 1) conserved Switch II residues t hat communicate the conformational state of the alpha subunit and 2) d ivergent residues that specify particular effecters and the nature of their modulation.