Ap. Uzerman et al., SITE-DIRECTED MUTAGENESIS OF THE HUMAN ADENOSINE A(2A) RECEPTOR - CRITICAL INVOLVEMENT OF GLU(13) IN AGONIST RECOGNITION, European journal of pharmacology, 310(2-3), 1996, pp. 269-272
A glutamic acid residue in the first transmembrane domain of the human
adenosine A(2A) receptor was mutated to glutamine. Radioligand bindin
g studies on COS-7 cell membranes expressing either the wild-type or t
he mutant receptor revealed that the affinity of the prototypic agonis
t CGS21680 phenyl]ethylamino]-5'-N-ethylcarboxamidoadenosine) for the
mutant receptor was 15-fold lower than for the wild-type receptor. Thi
s was confirmed in functional studies with intact cells. The EC(50) va
lues of CGS21680 for the stimulation of cAMP production differed in a
similar way. Antagonists of various chemical structure were equally ef
fective on both mutant and wild-type receptors, thus the mutation sele
ctively diminishes agonist affinity, We propose an indirect perturbati
on of the binding site, perhaps through a proton transfer mechanism as
suggested by molecular modelling.