Activation of the Rat P2X7 Receptor by Functionally Different ATP Activation Sites
The homotrimeric P2X7 receptor (P2X7R) contains three ATP<sup>4−</sup> binding sites in its ectodomain. Here, we investigated the role of individual ATP<sup>4−</sup> activation sites in rat P2X7R (rP2X7R) using trimeric concatemers consisting of either three wild-type subunit...
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2025-06-01
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| author | Fritz Markwardt Malte Berthold Sanaria Hawro Yakoob Günther Schmalzing |
| author_facet | Fritz Markwardt Malte Berthold Sanaria Hawro Yakoob Günther Schmalzing |
| author_sort | Fritz Markwardt |
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| description | The homotrimeric P2X7 receptor (P2X7R) contains three ATP<sup>4−</sup> binding sites in its ectodomain. Here, we investigated the role of individual ATP<sup>4−</sup> activation sites in rat P2X7R (rP2X7R) using trimeric concatemers consisting of either three wild-type subunits (7-7-7) or one to three subunits with ATP binding sites knocked out by the K64A mutation. Following expression in <i>Xenopus laevis</i> oocytes, ATP<sup>4−</sup>-elicited ion currents were recorded using the two-microelectrode voltage clamp technique. The 7-7-7 concatamer exhibited a biphasic ATP<sup>4−</sup> concentration dependence, best fit by the sum of two Hill functions, confirming the existence of functionally distinct ATP<sup>4−</sup> activation sites. The activation time course of the 7-7-7 was best approximated by the sum of a fast and a slow exponential saturating activation component. Similarly, deactivation exhibited both fast and slow exponential decay. Only one Hill function was required to best fit the ATP<sup>4−</sup> concentration dependence of concatamers with only two or one ATP<sup>4−</sup> binding sites, and their deactivation time courses largely lacked the slowly deactivating components. We conclude that the binding of one ATP<sup>4−</sup> is sufficient for partial activation of the rP2X7R and that allosteric effects occur when all three ATP<sup>4−</sup> binding sites are occupied, leading to distinct functional activation sites. |
| format | Article |
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| institution | Kabale University |
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| spelling | doaj-art-e4dd68415cb84daea1c6c1efee526af72025-08-20T03:27:28ZengMDPI AGCells2073-44092025-06-01141285510.3390/cells14120855Activation of the Rat P2X7 Receptor by Functionally Different ATP Activation SitesFritz Markwardt0Malte Berthold1Sanaria Hawro Yakoob2Günther Schmalzing3Julius-Bernstein-Institute of Physiology, Martin-Luther-University, D-06097 Halle, GermanyJulius-Bernstein-Institute of Physiology, Martin-Luther-University, D-06097 Halle, GermanyInstitute of Clinical Pharmacology, RWTH Aachen University, D-52074 Aachen, GermanyInstitute of Clinical Pharmacology, RWTH Aachen University, D-52074 Aachen, GermanyThe homotrimeric P2X7 receptor (P2X7R) contains three ATP<sup>4−</sup> binding sites in its ectodomain. Here, we investigated the role of individual ATP<sup>4−</sup> activation sites in rat P2X7R (rP2X7R) using trimeric concatemers consisting of either three wild-type subunits (7-7-7) or one to three subunits with ATP binding sites knocked out by the K64A mutation. Following expression in <i>Xenopus laevis</i> oocytes, ATP<sup>4−</sup>-elicited ion currents were recorded using the two-microelectrode voltage clamp technique. The 7-7-7 concatamer exhibited a biphasic ATP<sup>4−</sup> concentration dependence, best fit by the sum of two Hill functions, confirming the existence of functionally distinct ATP<sup>4−</sup> activation sites. The activation time course of the 7-7-7 was best approximated by the sum of a fast and a slow exponential saturating activation component. Similarly, deactivation exhibited both fast and slow exponential decay. Only one Hill function was required to best fit the ATP<sup>4−</sup> concentration dependence of concatamers with only two or one ATP<sup>4−</sup> binding sites, and their deactivation time courses largely lacked the slowly deactivating components. We conclude that the binding of one ATP<sup>4−</sup> is sufficient for partial activation of the rP2X7R and that allosteric effects occur when all three ATP<sup>4−</sup> binding sites are occupied, leading to distinct functional activation sites.https://www.mdpi.com/2073-4409/14/12/855purinoceptorP2X7activationstructurekineticsvoltage clamp |
| spellingShingle | Fritz Markwardt Malte Berthold Sanaria Hawro Yakoob Günther Schmalzing Activation of the Rat P2X7 Receptor by Functionally Different ATP Activation Sites Cells purinoceptor P2X7 activation structure kinetics voltage clamp |
| title | Activation of the Rat P2X7 Receptor by Functionally Different ATP Activation Sites |
| title_full | Activation of the Rat P2X7 Receptor by Functionally Different ATP Activation Sites |
| title_fullStr | Activation of the Rat P2X7 Receptor by Functionally Different ATP Activation Sites |
| title_full_unstemmed | Activation of the Rat P2X7 Receptor by Functionally Different ATP Activation Sites |
| title_short | Activation of the Rat P2X7 Receptor by Functionally Different ATP Activation Sites |
| title_sort | activation of the rat p2x7 receptor by functionally different atp activation sites |
| topic | purinoceptor P2X7 activation structure kinetics voltage clamp |
| url | https://www.mdpi.com/2073-4409/14/12/855 |
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