Figure 4.

Influence of rhodopsin on PDE activation by Gα_t and HD_t. The enzymatic activity of 0.2 nM PDE was determined with the indicated additions to the reactions. When present, rhodopsin was added at 1 μM, Gα_tGDP and Gα_tGTPγS were 125 nM, and HD_t was 0.5 μM. For nucleotide exchange, 5 μM GTPγS or 5 μM GDP were added with rhodopsin as indicated. The PDE reactions were conducted for 15 min. Error bars indicate ±SEM for values obtained in three independent experiments. The Gα_tGDP was purified as described (32), and its concentration was determined by rhodopsin-catalyzed GDP/[³⁵S]GTPγS exchange according to the modified method (19, 47).