Mutations of α1F45 residue of GABAA receptor loop G reveal its involvement in agonist binding and channel opening/closing transitions

• Published in: Biochemical pharmacology 2020-07, Vol.177, p.113917-113917, Article 113917
• Main Authors: Brodzki, Marek, Michałowski, Michał A., Gos, Michalina, Mozrzymas, Jerzy W.
• Format: Article
• Language: English
• Subjects: Binding; GABAA receptor; Gating; Loop G; Structure-function relationship
• ISSN: 0006-2952; 1873-2968
• DOI: 10.1016/j.bcp.2020.113917

[Display omitted] GABAA receptors (GABAARs) mediate inhibitory neurotransmission in the mammalian brain. Recently, numerous GABAAR static structures have been published, but the molecular mechanisms of receptor activation remain elusive. Loop G is a rigid β-strand belonging to an extensive β-sheet that spans the regions involved in GABA binding and the interdomain interface which is important in receptor gating. It has been reported that loop G participates in ligand binding and gating of GABAARs, however, it remains unclear which specific gating transitions are controlled by this loop. Analysis of macroscopic responses revealed that mutation at the α1F45 residue (loop G midpoint) resulted in slower macroscopic desensitization and accelerated deactivation. Single-channel analysis revealed that these mutations also affected open and closed times distributions and reduced open probability. Kinetic modeling demonstrated that mutations affected primarily channel opening/closing and ligand binding with a minor effect on preactivation. Thus, α1F45 residue, in spite of its localization close to binding site, affects late gating transitions. In silico structural analysis suggested an important role of α1F45 residue in loop G stability and rigidity as well as in general structure of the binding site. We propose that the rigid β-sheet comprising loop G is well suited for long range communication within GABAAR but this mechanism becomes impaired when α1F45 is mutated. In conclusion, we demonstrate that loop G is crucial in controlling both binding and gating of GABAARs. These data shed new light on GABAAR activation mechanism and may also be helpful in designing clinically relevant modulators.