Figure 4. Single turnover GTPase activities of Gtα* and Gtα*R238E. Effects of RGS9d and RGS16

Single turnover GTPase activities of Gtα* (A) and Gtα*R238E (B; 1 μM each) were measured in the presence of uROS membranes (5 μM rhodopsin) and 2 μM Gtβγ in the absence (rectangle) or presence of RGS9d (triangle) or RGS16 (down triangle). Reactions were started with the addition of 200 nM [γ-³²P]GTP (0.5 μCi), and free ³²P_i was measured by liquid scintillation. A: The k_cat values (s^-1) are 0.023±0.002 (rectangle), 0.093±0.006 (triangle), and 0.152±0.007 (down triangle). B: The k_cat values (s^-1) are 0.003 (rectangle), 0.004 (triangle), and 0.006 (down triangle). C: Kinetics of GTPγS binding to Gtα* (rectangle) and Gtα*R238E (triangle) under the conditions in (A,B) using 200 nM [³⁵S]GTPγS instead of [γ-³²P]GTP. The calculated k_app values (s^-1) were 0.16±0.01 (rectangle), and 0.14±0.01 (triangle).