A threonine residue in the seventh transmembrane domain of the human A1 adenosine receptor mediates specific agonist binding
The A1 adenosine receptor is a member of the seven-transmembrane G protein-coupled, receptor superfamily. This receptor binds the purine nucleoside adenosine with high affinity and inhibits the activity of adenylate cyclase. We have used site-directed mutagenesis and functional expression studies to examine the role of the threonine residue, located at position 277 in transmembrane domain VII of the human A1 receptor. Mutation of Thr-277 to either serine or alanine resulted in the expression of receptors that had essentially no change in binding affinity for the A1 selective antagonist 8-cyclo-pentyl-1,3-dipropylxanthine. Mutation of Thr-277 to serine resulted in modest (4.4-8.6-fold) but significant increases in the observed Ki values for three adenosine agonists, namely N-(1-methyl-2-phenethyl)adenosine (R-PIA or S-PIA) and 1-(6-amino-9H-purin-9-yl)-1-deoxy-N-ethyl-beta-L- ribofuranuronamide) (NECA). However, mutation of Thr-277 to alanine resulted in no significant changes in the Ki for R-PIA or S-PIA but did result in a highly significant 437-fold increase in the Ki for NECA. This demonstrates that the hydroxyl moiety of Thr-277 mediates agonist but not antagonist binding and, more specifically, that this residue forms a probable molecular contact site with the 5' substitution found in NECA.
|Authors||Townsend-Nicholson, A.;Schofield, P. R. :|
|Publisher Name||JOURNAL OF BIOLOGICAL CHEMISTRY|
|URL link to publisher's version||http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8300561|