GABAA receptor proline 273 at the interdomain interface of the β2 subunit regulates entry into desensitization and opening/closing transitions

Aims: GABA(A) receptors belong to Cys-loop ion channel family and mediate inhibition in the brain. Despite the abundance of structural data on receptor structure, the molecular scenarios of activation are unknown. In this study we investigated the role of a beta(2)P273 residue in channel gating transitions. This residue is located in a central position of the M2-M3 linker of the interdomain interface, expected to be predisposed to interact with another interfacial element, the beta 1-beta 2 loop of the extracellular side. The interactions occurring on this interface have been reported to couple agonist binding to channel gating. Main methods: We recorded micro- and macroscopic current responses of recombinant GABA(A) receptors mutated at the beta(2)P273 residue (to A, K, E) to saturating GABA. Electrophysiological data served as basis to kinetic modeling, used to decipher which gating transition were affected by mutations. Key findings: Mutations of this residue impaired macroscopic desensitization and accelerated current deactivation with P273E mutant showing greatest deviation from wild-type. Single-channel analysis revealed alterations mainly in short-lived shut times and shortening of openings, resulting in dramatic changes in intraburst open probability. Kinetic modeling indicated that beta(2)P273 mutants show diminished entry into desensitized and open states as well as faster channel closing transitions. Significance: In conclusion, we demonstrate that beta(2)P273 of the M2-M3 linker is a crucial element of the ECD-TMD interface regulating the receptor's ability to undergo late gating transitions. Henceforth, this region could be an important target for new pharmacological tools affecting GABA(A)R-mediated inhibition.

Automated summary

This study investigated the role of the beta(2)P273 residue in the gating transitions of GABA(A) receptors. The results showed that mutations of this residue impaired macroscopic desensitization and accelerated current deactivation. Furthermore, the mutations also affected the intraburst open probability of the receptors. The findings suggest that beta(2)P273 is a crucial element in regulating the receptor's ability to undergo late gating transitions and could be a potential target for pharmacological tools affecting GABA(A)R-mediated inhibition.