Atomic force microscopy imaging demonstrates that P2X2 receptors are trimers but that P2X6 receptor subunits do not oligomerize.
P2X receptors are cation-selective channels activated by extracellular ATP. The architecture of these receptors is still not completely clear. Here we have addressed this issue by both chemical cross-linking and direct imaging of individual receptors by atomic force microscopy (AFM). Cross-linking of the P2X(2) receptor produced higher order adducts, consistent with the presence of trimers. The mean molecular volume of the receptor determined by AFM (409 nm(3)) also points to a trimeric structure. P2X(2) receptors bearing His(6) epitope tags were incubated with anti-His(6) antibodies, and the resultant complexes were imaged by AFM. For receptors with two bound antibodies, the mean angle between the antibodies was 123 degrees , again indicating that the receptor is a trimer. In contrast, cross-linking of the P2X(6) receptor did not produce higher order adducts, and the mean molecular volume of the receptor was 145 nm(3). We conclude that P2X(2) receptors are trimers, whereas the P2X(6) receptor subunits do not form stable oligomers.
Barrera
Nelson P
NP
Department of Pharmacology, University of Cambridge, Tennis Court Road, Cambridge CB2 1PD, UK.
Ormond
Susan J
SJ
Henderson
Robert M
RM
Murrell-Lagnado
Ruth D
RD
Edwardson
J Michael
JM
D000818:Animals; D002460:Cell Line; D003902:Detergents / Q000494:pharmacology; D019281:Dimerization; D000939:Epitopes / Q000737:chemistry; D006639:Histidine / Q000737:chemistry; D006801:Humans; D015151:Immunoblotting; D018625:Microscopy, Atomic Force; D008954:Models, Biological; D016011:Normal Distribution; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D051381:Rats; D018048:Receptors, Purinergic P2 / Q000737:chemistry* / Q000502:physiology*; D058476:Receptors, Purinergic P2X2; D014162:Transfection
D003902:Detergents; D000939:Epitopes; C546401:P2RX2 protein, human; D018048:Receptors, Purinergic P2; D058476:Receptors, Purinergic P2X2; C110590:purinoceptor P2X6; D006639:Histidine