Table 5

K_D and B_max values for the binding of [³H]RX 821002, [³H]clonidine, and [³H]UK 14304 to membrane preparations of COS-7 cells expressing the α_2A-AR in either the absence or presence of various mutant G_αo proteins

G_αProtein	[³H]RX 821002		[³H]Clonidine		[³H]UK 14304
G_αProtein	K_D	B_max	K_D	B_max	K_D	B_max
	nM	pmol/mg protein	nM	pmol/mg protein	nM	pmol/mg protein
None	0.92 ± 0.09	30.70 ± 5.45	37.91 ± 2.16^5-a	5.04 ± 0.33^5-a	14.81 ± 2.80^5-a	4.12 ± 0.37^5-a
G_αo/GYGLY^5-150	0.92 ± 0.12	32.70 ± 6.90	1.41 ± 0.20	2.39 ± 0.38	0.33 ± 0.03	2.34 ± 0.30
G_αo/QYELL^5-160	1.14 ± 0.17	44.17 ± 14.1^5-a	26.10 ± 3.83^5-a	5.08 ± 1.34^5-a	3.07 ± 0.76^5-a	1.87 ± 0.47
G_αo/GYELL	0.91 ± 0.10	31.97 ± 8.47	42.39 ± 2.92^5-a	4.68 ± 0.72^5-a	9.76 ± 2.88^5-a	2.06 ± 0.69
G_αo/QYGLL	0.91 ± 0.06	29.42 ± 5.12	1.08 ± 0.12	2.83 ± 0.43	0.30 ± 0.03	2.48 ± 0.30
G_αo/GYGLL	0.88 ± 0.09	29.76 ± 4.66	2.15 ± 0.25	2.53 ± 0.39	0.38 ± 0.02	2.03 ± 0.21

Co-expression of α_2A-AR and either empty plasmid or respective mutant G_αo protein was performed as described under Experimental Procedures. All conditions were treated with PTX (20 ng/ml). The equilibrium dissociation constant (K_D, nM) and maximal radioligand binding capacity (B_max, pmol/mg of protein) were determined for each condition, as described under Experimental Procedures, according to a monophasic Scatchard analysis. Data represent mean values ± S.E.M. of four independent transfection experiments, each performed in duplicate. The bold amino acids correspond to those that are different between G_αo/s (G_αo/QYELL) and the various mutant G_αo proteins. Statistical analysis was performed on ligand's K_D andB_max values between G_αo/GYGLY and the other mutant G_αo proteins or empty plasmid.

↵5-150 , G_αoCys³⁵¹Tyr protein;
↵5-160 , G_αo/s protein.
↵5-a P < 0.05.