3 DISCRETE REGIONS OF MAMMALIAN ADENYLYL-CYCLASE FORM A SITE FOR G(S-ALPHA) ACTIVATION

The interaction between the alpha subunit of G protein G(s) (G(s alpha )) and the two cytoplasmic domains of adenylyl cyclase (C-1 and C-2) i s a key step in the stimulation of cAMP synthesis by hormones, Mutatio nal analysis reveals that three discrete regions in the primary sequen ce of adenylyl cyclase affect the EC50 values for G(s alpha) activatio n and thus are the affinity determinants of G(s alpha). Based on the t hree-dimensional structure of C-2 forskolin dimer, these three regions (C-2, alpha 2, C-2 alpha 3/beta 4, and C-1 beta 1) are close together and form a negatively charged and hydrophobic groove the width of an alpha helix that can accommodate the positively charged adenylyl cycla se binding region of G(s alpha). Two mutations in the C-2 alpha 3/beta 4 region decrease the V-max values of G(s alpha) activation without a n increase in the EC50 values, Since these three regions are distal to the catalytic site, the likely mechanism for G(s alpha) activation is to modulate the structure of the active site by controlling the orien tation of the C-2 alpha 2 and alpha 3/beta 4 structures.