Φ value analysis underscores strong functional and structural compactness of the GABAA receptor

• 成果类型: Article
• 署名作者: Michaowski, Michal A.; Terejko, Katarzyna; Gos, Michalina; Izykowska, Ilona; Czyzewska, Marta M.; Kopotowski, Karol; Kaczor, Przemysaw T.; Brzostowicz, Aleksandra; Puzek, Estera; Migdaek, Monika; Mozrzymas, Jerzy W.
• 署名单位: Wroclaw Medical University; University of Wroclaw; Wroclaw University of Environmental & Life Sciences; Polish Academy of Sciences; Hirszfeld Institute of Immunology & Experimental Therapy of the Polish Academy of Sciences
• 刊物名称: PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
• ISSN/ISSBN: 0027-12646
• DOI: 10.1073/pnas.2512278122
• 发表日期: 2025-09-16
• 关键词: benzodiazepine-receptor; crystal-structures; binding-site; channel; acetylcholine; modulation; mechanism; desensitization; diazepam; subunits

摘要

gamma- aminobutyric acid type A receptor (GABAAR) is a pentameric ligand-gated ion channel that plays a crucial role in inhibition in the adult brain. Structural and electrophysiological studies have provided numerous insights into the receptor's functioning but the complete molecular mechanism of GABAAR action remains elusive. Herein, we used high-resolution single-channel recording to analyze point-mutated alpha 1 beta 2 gamma 2 receptors and applied so called Phi value (REFER, rate-equilibrium free energy relationship) analysis which allows to infer the order of domains engagement during activation, offering a complementary dynamic insight into the receptor's function. As anticipated, point mutations at the orthosteric binding sites reduced GABA binding affinity, with the magnitude of this effect diminishing progressively with increasing distance from the binding site. On the contrary, mutations located all over the macromolecule's structure, e.g., in peripheral top position, extracellular/transmembrane interface, and channel pore, affected the receptor with a clear tendency to influence entire gating including opening/closing, preactivation, and desensitization with no clear correlation with the distance to the channel gate. Interestingly, the calculated Phi values for the GABAAR showed a relatively narrow range (0.42 to 0.81), suggesting that the conformational transitions are highly synchronized and coordinated by a global network of interactions. This prediction is also in agreement with observation that, typically, single GABAAR residue mutation alters the kinetics of not just one but many (often all) conformational transitions. We thus propose a dictum reflecting modus operandi of the GABAAR: Binding is local and gating is global. In conclusion, our analysis indicates that GABAAR shows particularly strong functional compactness manifested as global gating mechanisms and high allostery.