Influence of GABAA Receptor α Subunit Isoforms on the Benzodiazepine Binding Site

Abstract

Classical benzodiazepines, such as diazepam, interact with αxβ2γ2 GABAA receptors, x = 1, 2, 3, 5 and modulate their function. Modulation of different receptor isoforms probably results in selective behavioural effects as sedation and anxiolysis. Knowledge of differences in the structure of the binding pocket in different receptor isoforms is of interest for the generation of isoform-specific ligands. We studied here the interaction of the covalently reacting diazepam analogue 3-NCS with α1S204Cβ2γ2, α1S205Cβ2γ2 and α1T206Cβ2γ2 and with receptors containing the homologous mutations in α2β2γ2, α3β2γ2, α5β1/2γ2 and α6β2γ2. The interaction was studied using radioactive ligand binding and at the functional level using electrophysiological techniques. Both strategies gave overlapping results. Our data allow conclusions about the relative apposition of α1S204Cβ2γ2, α1S205Cβ2γ2 and α1T206Cβ2γ2 and homologous positions in α2, α3, α5 and α6 with C-atom adjacent to the keto-group in diazepam. Together with similar data on the C-atom carrying Cl in diazepam, they indicate that the architecture of the binding site for benzodiazepines differs in each GABAA receptor isoform α1β2γ2, α2β2γ2, α3β2γ2, α5β1/2γ2 and α6β2γ2.

A Benzodiazepine Ligand with Improved GABAA Receptor α5-Subunit Selectivity Driven by Interactions with Loop C

“…Loop C, on the other hand, has long been known to be important for ligand binding, since it is more variable than the other loops (Michalowski et al, 2017). α1-serine 206 (neighboring the serine mutated in our study) seems to interact physically with diazepam (Luscher et al, 2012). Both α1-serine 206 as well as α1-tyrosine 209 are important in determining the binding affinities for ligands of the benzodiazepine binding (Buhr et al, 1997) and S206 has been shown to influence the efficacy of midazolam to modulate GABA induced currents (Moody and Jenkins, 2018) as well as the affinities for β-carboline binding (Derry et al, 2004).…”

Metal-assisted synthesis of unsymmetrical magnolol and honokiol analogs and their biological assessment as GABAA receptor ligands

“…Compounds were applied for 20 s. None of the compounds could open the channel without GABA, so they all are only modulators of the channel. For details on the electrophysiological recordings see Lüscher et al 23 …”

The role of loops B and C in determining the potentiation of GABAA receptors by midazolam

“…Previous studies found that the α6(Asn204) and α4(Ile203) residues (both homologous to human α1(Ser206)) were important for distinguishing the binding of negative benzodiazepines . Ser206 also physically interacts with diazepam in α1, α2 and α5, suggesting a critical role in benzodiazepine action . However, a neighboring mutation, homologous to α1(T207C), specifically altered benzodiazepine efficacy and not binding .…”

“…and α5, suggesting a critical role in benzodiazepine action. 37 However, a neighboring mutation, homologous to α1(T207C), specifically altered benzodiazepine efficacy and not binding. 12 We propose that the homologous Ser206 in loop C may provide an important point of contact between the ligand and benzodiazepine site that affects the coupling of the benzodiazepine site to GABA activation, thereby affecting the benzodiazepine's efficacy.…”