Dynamic Interactions in the Human β₁AR Signalling Complex with Mini-G_s Revealed by NMR

A wealth of insights into the mechanisms of GPCRs have been gained from biophysical studies of thermostabilized β₁ adrenergic receptors (β₁AR) from turkey. Here we investigate a stabilised variant of the pharmacologically more relevant human β₁AR and present initial insights into the active signalling complex with the G protein surrogate mini-G_s. Compared to the avian receptor, the human β₁AR construct exhibits greater conformational flexibility and more readily transitions between inactive and pre-active states; however, the transition to the fully active, open state remains slow. Interestingly, in contrast, the bound mini-G_s protein exhibits much faster dynamics in the ternary complex. For the receptor in the signalling complex, we reveal at least two interconverting states of intracellular loop 2 and the extracellular end of transmembrane helix 1. Additionally, we demonstrate that intracellular loop 3 contributes to mini-G_s binding. This human β₁AR construct provides a valuable platform for biophysical studies at the atomic level towards understanding the behaviour of the human receptor.