GABA type A receptor plays a key role in inhibitory signaling in the adult central nervous system. This receptor can be modulated by protons but the underlying molecular mechanisms have not been fully explored. To find possible pH-sensor residues, a comparative study for proton-activated GLIC channel and α1β2γ2 GABA receptor was performed and pK 's of respective residues were estimated by numerical algorithms which consider local interactions. β E155, located at the GABA binding site, showed pKa values close to physiological values and dependence on the receptor state and ligation, suggesting a role in modulation by pH. To validate this prediction, pH sensitivity of current responses to GABA was investigated using patch-clamp technique for WT and mutated (β2E155[C, S, Q, L]) GABA receptors. Cysteine mutation preserved pH sensitivity. However, for remaining mutants, the sensitivity to acidification (pH = 6.0) was reduced becoming not statistically significant. The effect of alkaline pH (8.0) was maintained for all mutants with exception for β2E155L for which it was nearly abolished. To further explore the impact of considered mutations, molecular docking was performed which indicated that pH modulation is probably affected by interplay between binding site residues, zwitterion GABA and protons. These data, altogether, indicate that mutation of β2E155 to hydrophobic residue (L) maximally impaired pH modulation while for polar substitutions the effect was smaller. In conclusion, our data provide evidence that a key binding site residue β2E155 plays an important role in proton sensitivity of GABA receptor.
Keywords: GABA; Gating; Ion ihannel; Modulation; Receptor; pH.
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