Mutations in the palm domain disrupt modulation of acid-sensing ion channel 1a currents by neuropeptides

Abstract

Modulation by neuropeptides enhances several functions of acid-sensing ion channels (ASICs), such as pain sensation and acid-induced neuronal injury. The acid-induced opening of ASICs is transient, because of a rapid desensitization. Neuropeptides containing an Arg-Phe-amide motif affect ASIC desensitization and allow continuous activity of ASICs. In spite of the importance of the sustained ASIC activity during prolonged acidification, the molecular mechanisms of ASIC modulation by neuropeptides is only poorly understood. To identify the FRRFa (Phe-Arg-Arg-Phe-amide) binding site on ASIC1a, we carried out an in silico docking analysis and verified functionally the docking predictions. The docking experiments indicated three possible binding pockets, located (1) in the acidic pocket between the thumb, finger, β-ball and palm domains, (2) in a pocket at the bottom of the thumb domain, and (3) in the central vestibule along with the connected side cavities. Functional measurements of mutant ASIC1a confirmed the importance of residues of the lower palm, which encloses the central vestibule and its side cavities, for the FRRFa effects. The combined docking and functional experiments strongly suggest that FRRFa binds to the central vestibule and its side cavities to change ASIC desensitization.

Mechanism and site of action of big dynorphin on ASIC1a

“…Together, this argues for an extended interaction surface on ASIC1a, likely extending from the peripheral thumb domain around the α5 helix and into the acidic pocket. While the location of the BigDyn binding site is distinct from that suggested for RFamide neuropeptides on ASICs (41,42), it does overlap with the binding site for PcTx1 (10,25,26,31). This is consistent with the finding that RFamides do not functionally compete with BigDyn or PcTx1 (10,43).…”

Sevanol and Its Analogues: Chemical Synthesis, Biological Effects and Molecular Docking

“…The peptides exhibit this effect both in the case of desensitization following the activation of the channels and in the case of steady-state desensitization, when the channels cease to respond to the acid stimulus bypassing the activation stage [ 35 , 36 ]. Using molecular docking and site-directed mutagenesis, it has been shown that RF-amide peptides are most likely to bind to ASIC1a in the central vestibule of the chicken channel [ 37 ]. We examined the inhibitory effect of sevanol on the ASIC1a channel activated by pH 5.5 in the presence of 200 μM FRRFa peptide (Phe-Arg-Arg-Phe-amide), and showed that sevanol and the peptide compete for the binding site.…”

“…Our electrophysiological experiments show that sevanol and FRRFa compete to a small extent for the binding site, and this fact is confirmed by the docking results. Recently, FRRFa has been predicted to bind in the side cavities of the vestibule, coordinated by three glutamic acid and one valine residues that have been confirmed to be important for this binding [37]. Sevanol, as a highly acidic molecule, is unlikely to occupy the same site due to a presumed strong electrostatic repulsion.…”

“…The yellow sticks represent 20 sevanol docking solutions inside the central vestibule. Magenta tubes depict the docking results for the FRRFa peptide (obtained from[37]), revealing three probable binding sites per subunit (the so-called acidic pocket, the pocket at the bottom of the thumb domain, and the central vestibule side cavities). (B) Close-up view of (A) with the central vestibule cavity shown as a semi-transparent yellow surface.…”

Acid-Sensing Ion Channels

“…FMRFamide has also been shown to modulate the function of ASIC1 and ASIC3 by potentiating their responses to acidification. The action on ASICs of FMRFamide and related neuropeptides involves amino acid residues in the palm and a linker between the palm and the thumb (Vick and Askwith 2015;Bargeton et al 2019). FMRFamide and related neuropeptides increase ASIC activity in several ways, including an acidic shift of the pH dependence of desensitization, a slowing of the current decay, and the generation of a sustained current (Fig.…”