Three discrete regions of mammalian adenylyl cyclase form a site for activation

Abstract

The interaction between the α subunit of G protein G(s) (G(sα)) and the two cytoplasmic domains of adenylyl cyclase (C1 and C2) is a key step in the stimulation of cAMP synthesis by hormones. Mutational analysis reveals that three discrete regions in the primary sequence of adenylyl cyclase affect the EC50 values for G(sα) activation and thus are the affinity determinants of G(sα). Based on the three-dimensional structure of C2·forskolin dimer, these three regions (C2 α2, C2 α3/β4, and C1 β1) are close together and form a negatively charged and hydrophobic groove the width of an α helix that can accommodate the positively charged adenylyl cyclase binding region of G(sα). Two mutations in the C2 α3/β4 region decrease the V(max) values of G(sα) activation without an increase in the EC50 values. Since these three regions are distal to the catalytic site, the likely mechanism for G(sα) activation is to modulate the structure of the active site by controlling the orientation of the C2 α2 and α3/β4 structures.