Identification of Cinnamic Acid Derivatives As Novel Antagonists of the Prokaryotic Proton-Gated Ion Channel GLIC

Abstract

Pentameric ligand gated ion channels (pLGICs) mediate signal transduction. The binding of an extracellular ligand is coupled to the transmembrane channel opening. So far, all known agonists bind at the interface between subunits in a topologically conserved "orthosteric site" whose amino acid composition defines the pharmacological specificity of pLGIC subtypes. A striking exception is the bacterial proton-activated GLIC protein, exhibiting an uncommon orthosteric binding site in terms of sequence and local architecture. Among a library of Gloeobacter violaceus metabolites, we identified a series of cinnamic acid derivatives, which antagonize the GLIC proton-elicited response. Structure-activity analysis shows a key contribution of the carboxylate moiety to GLIC inhibition. Molecular docking coupled to site-directed mutagenesis support that the binding pocket is located below the classical orthosteric site. These antagonists provide new tools to modulate conformation of GLIC, currently used as a prototypic pLGIC, and opens new avenues to study the signal transduction mechanism.

Structural evidence for the binding of monocarboxylates and dicarboxylates at pharmacologically relevant extracellular sites of a pentameric ligand-gated ion channel

“…This latter receptor from a tubeworm symbiont bacterium, named sTeLIC, is strongly potentiated by a cinnamic acid derivative that binds at a specific vestibular site of the extracellular domain located behind the common pLGIC orthosteric site (Hu et al, 2018). Interestingly, GLIC is also modulated by cinnamic acid derivatives as well as other carboxylic acid derivatives such as caffeic and crotonic acids, which display consistent inhibition of GLIC currents (Prevost et al, 2013;Alqazzaz et al, 2016). While these compounds have been suggested to bind at the pLGIC orthosteric site in GLIC by docking studies, no experimental data are yet available to support this hypothesis.…”

“…These sites were called the 'intrasubunit' and 'intersubunit' acetate sites, respectively, according to their localization within or between the receptor subunits. Our rationale for this study was to provide experimental evidence for the binding of carboxylate derivatives other than acetate to GLIC, especially those for which the modulation potency has previously been described, namely caffeic acid and crotonic acid (Alqazzaz et al, 2016;Prevost et al, 2013). While all of our attempts to crystallize GLIC with caffeic acid yielded only poorly diffracting crystals, we managed to solve the structure of GLIC in complex with crotonic acid, as well as with a series of other carboxylate derivatives (Table 1).…”

Isolation and identification of a growth inhibitory substance from Heliotropium indicum L.

“…The substance was characterized as methyl caffeate (Fig. 2) by comparison of the spectroscopic data with previously reported in the literature [23,32].…”

Plant growth inhibitory activity and active substances with allelopathic potential of cogongrass (Imperata cylindrica) rhizome

“…The substance was identified as “methyl caffeate” (Fig. ) by comparing it with previous data (Prevost et al ).…”

“…A series of the bioassay‐guided separations of the extracts of the cogongrass rhizomes led to the isolation of the four growth inhibitory substances (substances 1, 2, 3 and 4). Substance 1, 2, 3 and 4 were identified as abscisic acid, 5‐methoxyflavone, 5,2′‐dimethoxyflavone and methyl caffeate, respectively, by comparison of HRESI‐MS, 1 H‐NMR and 13 C‐NMR spectra data with that in the literature (Ferreres et al ; Lee et al ; Prevost et al ) (Fig. ).…”