Molecular Mechanism of the β3AR Agonist Activity of a β‐Blocker

Development of subtype‐selective drugs for G protein‐coupled receptors poses a significant challenge due to high similarity between subtypes, as exemplified by the three β‐adrenergic receptors (βARs). The β 3 AR agonists show promise for treating the overactive bladder or preterm birth, but their potential is hindered by off‐target activation of β 1 AR and β 2 AR. Interestingly, several β‐blockers, which are antagonists of the β 1 ARs and β 2 ARs, have been reported to exhibit agonist activity at the β 3 AR. However, the molecular mechanism remains elusive. Understanding the underlying mechanism should facilitate the development of β 3 AR agonists with improved selectivity and reduced off‐target effects. In this work, we determined the structures of human β 3 AR in complex with the endogenous agonist epinephrine or with a synthetic β 3 AR agonist carazolol, which is also a high‐affinity β‐blocker. Structure comparison, mutagenesis studies and molecular dynamics simulations revealed that the differences on the flexibility of D 3.32 directly contribute to carazolol's distinct activities as an antagonist for the β 2 AR and an agonist for the β 3 AR. The process is also indirectly influenced by the extracellular loops (ECL), especially ECL1. Taken together, these results provide key guidance for development of selective β 3 AR agonists, paving the way for new therapeutic opportunities.