TOPOLOGICAL ANALYSIS OF THE ATP-GATED IONOTROPHIC P2X(2) RECEPTOR SUBUNIT

We investigated the transmembrane topology of the P2X(2) receptor subu nit expressed in HEK 293 cells, Initial studies using two P2X subunits expressed in tandem indicated that the amino-and carboxy-termini are on the same side of the membrane, Immunofluorescence studies showed th e cytoplasmic orientation of the amino-and carboxy-termini. Finally, N -glycosylation scanning mutagenesis revealed that reporter sites inser ted into the central loop, but not those in the amino-or carboxy-termi nal regions, were glycosylated, thus suggesting an extracellular place ment for that domain, Our results support a two-transmembrane arrangem ent for P2X receptors with intracellular amino-and carboxy-termini. (C ) 1998 Federation of European Biochemical Societies.