Abstract
P2X receptors are a family of seven ligand-gated ion channels (P2X1-P2X7) that open in the presence of ATP. We used alanine-scanning mutagenesis and patch clamp photometry to study the role of the first transmembrane domain of the rat P2X2 receptor in cation permeability and flux. Three alanine-substituted mutants did not respond to ATP, and 19 of the 22 functional receptors resembled the wild-type receptor with regard to the fraction of the total ATP-gated current carried by calcium or the permeability of calcium relative to cesium. The remaining three mutants showed modest changes in calcium dynamics. Two of these occurred at sites (Gly30 and Phe44) that are unlikely to interact with permeating cations in a meaningful way. The third was a conserved tyrosine (Tyr43) that may form an inter-pore binding site for calcium. The data suggest that, with the possible exception of Tyr43, the first transmembrane domain contributes little to the permeation properties of the P2X2 receptor.
Highlights
In this study, we consider the role of TM1 in the cation permeability and flux of the P2X2 receptor
We found that systematic mutagenesis produced no explainable changes in either Pf% or PCa/PCs, suggesting that TM1 plays a limited role in controlling cation permeability and flux through the P2X2 pore
Alanine-scanning Mutagenesis—Activation of wild-type P2X2 receptors with 3 M ATP evoked a Pf% of 6.6 Ϯ 0.2% (n ϭ 22; Fig. 1A), a value in keeping with published results [10, 12]
Summary
We consider the role of TM1 in the cation permeability and flux of the P2X2 receptor. There is general agreement that TM2 contributes, because published data show that some of its side chains form a water-accessible surface [6, 7] that participates in control of cation conductance [8], permeability [9], and flux [10]. We use a combination of whole-cell voltage clamp technology, site-directed mutagenesis, and fluorescence microscopy to study the role of TM1 in Ca2ϩ permeability and flux. We found that systematic mutagenesis produced no explainable changes in either Pf% or PCa/PCs, suggesting that TM1 plays a limited role in controlling cation permeability and flux through the P2X2 pore
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
