Structural determinants for interaction with three different effectors on the G protein β subunit

Abstract

In the yeast two-hybrid system, a 100-residue fragment (β1A) from the N terminus of the β1 subunit interacts with domains specific to adenylyl cyclase 2 (AC2), the muscarinic atrial potassium channel (GIRK1), and phospholipase C-β2 (PLC-β2). Based on the crystal structure of the G protein, β1A is composed of an N-terminal α helix, a loop, and five β strands in which the C-terminal four β strands form a β sheet, the first of seven sheets that make up the propeller structure of the β subunit. A mutant of β1A (L4P, L7P, and L14P), in which the α helix was potentially destroyed, interacted poorly with the G protein γ subunit but effectively with domains of AC2, GIRK1, and PLC-β2. In contrast, another mutant of β1A (S72A, D76A, and W82A), in which a network of hydrogen bonds was disrupted, interacted poorly with GIRK1 and PLC-β2 domains, but effectively with the γ subunit and the AC2 domain. These results suggest that the proper folding of the first five β strands in the G protein β subunit is a requirement for appropriately positioning residues that interact with GIRK1 and PLC-β2. Furthermore, since mutations that potentially disrupted the folding of these β strands did not affect interaction with AC2, the structural determinants on the G protein β subunit for interaction with various effectors may be different.