Abstract
Gating of the ATP-activated channel P2X2 has been shown to be dependent not only on [ATP] but also on membrane voltage, despite the absence of a canonical voltage-sensor domain. We aimed to investigate the structural rearrangements of rat P2X2 during ATP- and voltage-dependent gating, using a voltage-clamp fluorometry technique. We observed fast and linearly voltage-dependent fluorescence intensity (F) changes at Ala337 and Ile341 in the TM2 domain, which could be due to the electrochromic effect, reflecting the presence of a converged electric field. We also observed slow and voltage-dependent F changes at Ala337, which reflect structural rearrangements. Furthermore, we determined that the interaction between Ala337 in TM2 and Phe44 in TM1, which are in close proximity in the ATP-bound open state, is critical for activation. Taking these results together, we propose that the voltage dependence of the interaction within the converged electric field underlies the voltage-dependent gating.
Highlights
30 IntroductionP2X2 is a member of the P2X receptor family, a ligand-gated cation channel which opens upon the binding of extracellular ATP (Brake et al, 1994; Valera et al, 1994)
The present study aims at defining the roles of the TM domains of the P2X2 receptor in complex gating by [ATP] and voltage, using voltage-clamp fluorometry (VCF) with a genetically incorporated fluorescent unnatural amino acid (fUAA) probe
The change could be well interpreted to be due to an electrochromic effect, indicating that there is an electric field convergence at both positions, which are located adjacent to each other
Summary
30 IntroductionP2X2 is a member of the P2X receptor family, a ligand-gated cation channel which opens upon the binding of extracellular ATP (Brake et al, 1994; Valera et al, 1994). P2X2 in the cochlea is involved in adaptation to elevated sound levels (Housley et al, 2013)
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