Activation of the G(s) heterotrimer monitored in living cells with novel G(s) biosensors

Abstract

Our understanding of conformational changes associated with receptor activation has been advanced profoundly by the crystal structure of agonist‐bound β2 adrenergic receptor (β2AR) coupled to G(s). However, there is uncertainty regarding the extent to which conformational changes in G protein are conserved in living cells as well as across different receptors.We set out to understand the movement of the G(s) heterotrimer after activation of β2AR in intact cells. Using bioluminescence resonance energy transfer, we made observations on movement within the G protein and between the receptor and the G protein. Using a library of novel G(s) biosensors with either luciferase or GFP inserted at various positions throughout the structure, we studied conformational changes in cells and compared the results to the crystal structures of the inactive and active conformations. We also studied conformational changes in G(s) protein induced by the D1 dopamine receptor and A2A adenosine receptor.Our analysis using these G(s) biosensors suggests that conformational changes within the G(s) heterotrimer are largely conserved among different G(s)‐coupling receptors. Using this set of G(s) biosensors, partiality of agonists can be studied in relation to structural changes and subsequent effector activation. Thus, these biosensors represent a novel pharmacological tool to study structure‐function relationships.