Roles of individual N-glycans for ATP potency and expression of the rat P2X1 receptor

  • P2X1 receptor subunits assemble in the ER of Xenopus oocytes to homotrimers that appear as ATP-gated cation channels at the cell surface. Here we address the extent to which N-glycosylation contributes to assembly, surface appearance, and ligand recognition of P2X1receptors. SDS-polyacrylamide gel electrophoresis (PAGE) analysis of glycan minus mutants carrying Gln instead of Asn at five individual NXT/S sequons reveals that Asn284 remains unused because of a proline in the +4 position. The four other sites (Asn153, Asn184, Asn210, and Asn300) carryN-glycans, but solely Asn300 located only eight residues upstream of the predicted reentry loop of P2X1acquires complex-type carbohydrates. Like parent P2X1, glycan minus mutants migrate as homotrimers when resolved by blue native PAGE. Recording of ATP-gated currents reveals that elimination of Asn153 or Asn210 diminishes or increases functional expression levels, respectively. In addition, elimination of Asn210 causes a 3-fold reduction of the potency for ATP. If three or all four N-glycosylation sites are simultaneously eliminated, formation of P2X1 receptors is severely impaired or abolished, respectively. We conclude that at least oneN-glycan per subunit of either position is absolutely required for the formation of P2X1 receptors and that individual N-glycans possess marked positional effects on expression levels (Asn154, Asn210) and ATP potency (Asn210).

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Author:Jürgen RettingerGND, Armaz AschrafiORCiD, Günther SchmalzingORCiDGND
Pubmed Id:
Parent Title (English):The journal of biological chemistry
Publisher:American Society for Biochemistry and Molecular Biology Publications
Place of publication:Bethesda, Md
Document Type:Article
Date of Publication (online):2021/01/04
Year of first Publication:2000
Publishing Institution:Universitätsbibliothek Johann Christian Senckenberg
Release Date:2023/11/08
Page Number:6
First Page:33542
Last Page:33547
Institutes:Biochemie, Chemie und Pharmazie
Dewey Decimal Classification:6 Technik, Medizin, angewandte Wissenschaften / 61 Medizin und Gesundheit / 610 Medizin und Gesundheit
Licence (German):License LogoCreative Commons - CC BY - Namensnennung 4.0 International