Effects of magnesium on cyclic GMP hydrolysis by the bovine retinal rod cyclic GMP phosphodiesterase

Knowledge of the kinetics of the rod cyclic GMP phosphodiesterase is essential for understanding the kinetics and gain of the light response. Therefore, the interactions between Mg²⁺, cyclic GMP, and purified, trypsin-activated bovine rod cyclic GMP phosphodiesterase (EC 3.1.4.17) were examined. The effects of Mg²⁺ and of cyclic GMP on the rod phosphodiesterase activity were mutually concentration-dependent. Formation of a free Mg-cyclic GMP complex is unlikely due to its high dissociation constant (K(d) = 19 mM). Plots of 1/velocity versus 1/[cyclic GMP] as a function of [Mg²⁺] and 1/velocity versus 1/[Mg²⁺] as a function of [cyclic GMP] intersected to the left of the 1/velocity axis. This is consistent with the formation of a ternary complex between the phosphodiesterase, Mg²⁺, and cyclic GMP. A competitive inhibitor of the phosphodiesterase relative to cyclic GMP, 3-isobutyl-1-methylxanthine, non-competitively inhibited the enzyme relative to Mg²⁺. Pb²⁺, a competitive inhibitor of the phosphodiesterase relative to Mg²⁺ non-competitively inhibited the enzyme relative to cyclic GMP. Collectively these results are suggestive of a rapid equilibrium random binding order of Mg²⁺ and cyclic GMP to the rod phosphodiesterase.