Amino Acid Residues in the P2X7 Receptor that Mediate Differential Sensitivity to ATP and BzATP

Agonist properties of the P2X₇ receptor (P2X₇R) differ strikingly from other P2X receptors in two main ways: high concentrations of ATP (> 100 μM) are required to activate the receptor, and the ATP analog 2′,3′-O-(4-benzoyl-benzoyl)ATP (BzATP) is both more potent than ATP and evokes a higher maximum current. However, there are striking species differences in these properties. We sought to exploit the large differences in ATP and BzATP responses between rat and mouse P2X₇R to delineate regions or specific residues that may be responsible for the unique actions of these agonists at the P2X₇R. We measured membrane currents in response to ATP and BzATP at wild-type rat and mouse P2X₇R, at chimeric P2X₇Rs, and at mouse P2X₇Rs bearing point mutations. Wild-type rat P2X₇R was 10 times more sensitive to ATP and 100 times more sensitive to BzATP than wild-type mouse P2X₇R. We found that agonist EC₅₀ values were determined solely by the ectodomain of the P2X₇R. Two segments (residues 115-136 and 282-288), when transposed together, converted mouse sensitivities to those of rat. Point mutations through these regions revealed a single residue, asparagine²⁸⁴, in the rat P2X₇R that fully accounted for the 10-fold difference in ATP sensitivity, whereas the 100-fold difference in BzATP sensitivity required the transfer of both Lys¹²⁷ and Asn²⁸⁴ from rat to mouse. Thus, single amino acid differences between species can account for large changes in agonist effectiveness and differentiate between the two widely used agonists at P2X₇ receptors.