TABLE 1. Ligand binding properties of wild-type, E¹⁶¹A, and E¹⁶⁹Q human A_2a, receptors.

Agonist and antagonist binding affinities (K_i, values) were determined in [³H]CGS 21680 (15 nm) competition binding studies using membrane homogenates prepared from transiently transfected COS-7 cells, as described in Experimental Procedures. K_i, values were calculated from IC₅₀ value with the Kaleidagraph program. Approximately 15 μg of membrane protein/tube were used. The following K_d (nm) and B_max values for [³H]CGS 21680 (pmol/mg protein, in parentheses) were determined: wild-type, 22.3 ± 4.6 (15.5 ± 0.1); E¹⁶¹A, 41.7 ± 9.2 (17.0 ± 2.7); and E¹⁶⁹Q, 57.0 ± 1.8 (9.26 ± 0.37). Values are mean ± standard error of two or three independent experiments, each performed in duplicate.

ligand	Position of substitution (agonists)	Ki
		Wild-type	E161A	E160Q
		nm
Agonist  2-CADO	2	144 ± 33	221 ± 71	946 ± 20
DPMA	N6	36.3 ± 1.2	61.2 ± 0.2	1.62 ± 0.94
R-N6-Phenyiisopropyladenosine	N6	158 ± 19	264 ± 21	33.9 ± 2.8
Agonist  IB-MECA	N6,5′	456 ± 56	433 ±13	53.4 ± 2.6
NECA	5′	11.4 ± 4.0	23.9 ± 0.2	253 ± 40
Antagonist  CGS 15943		1.71 ± 0.28	0.287 ± 0.16	2.26 ± 1.4
XAC		6.98 ± 1.6	7.19 ± 1.70	11.5 ± 2.8