Molecular determinants involved in the allosteric control of agonist affinity in the GABAB receptor by the GABAB2 subunit

TL;DR

This study uncovers how the GABAB2 subunit's Venus flytrap domain regulates agonist affinity in the GABAB receptor by modulating domain interactions, revealing new insights into receptor allosteric control mechanisms.

Contribution

It demonstrates that the GABAB2 VFT domain directly influences GABAB1 agonist affinity and prevents inhibitory domain interactions, providing a novel understanding of allosteric regulation in GABAB receptors.

Findings

GABAB2 VFT forms hetero-oligomers with GABAB1 VFT.

GABAB2 VFT increases GABAB1 agonist affinity.

GABAB2 VFT prevents inhibitory VFT-HD interactions in GABAB1.

Abstract

The gamma-aminobutyric acid type B (GABAB) receptor is an allosteric complex made of two subunits, GABAB1 (GB1) and GABAB2 (GB2). Both subunits are composed of an extracellular Venus flytrap domain (VFT) and a heptahelical domain (HD). GB1 binds GABA, and GB2 plays a major role in G-protein activation as well as in the high agonist affinity state of GB1. How agonist affinity in GB1 is regulated in the receptor remains unknown. Here, we demonstrate that GB2 VFT is a major molecular determinant involved in this control. We show that isolated versions of GB1 and GB2 VFTs in the absence of the HD and C-terminal tail can form hetero-oligomers as shown by time-resolved fluorescence resonance energy transfer (based on HTRF technology). GB2 VFT and its association with GB1 VFT controlled agonist affinity in GB1 in two ways. First, GB2 VFT exerted a direct action on GB1 VFT, as it slightly increased agonist affinity in isolated GB1 VFT. Second and most importantly, GB2 VFT prevented inhibitory interaction between the two main domains (VFT and HD) of GB1. According to this model, we propose that GB1 HD prevents the possible natural closure of GB1 VFT. In contrast, GB2 VFT facilitates this closure. Finally, such inhibitory contacts between HD and VFT in GB1 could be similar to those important to maintain the inactive state of the receptor.