Kinetic and thermodynamic studies on nitrobenzylthioinosine binding to the nucleoside transporter of Chinese hamster ovary cells.

The binding of [G-3H]nitrobenzylthioinosine to intact Chinese hamster ovary cells has been studied kinetically and thermodynamically. The association of nitrobenzylthioinosine with cells is a second-order process which proceeds at 24 degrees C with a rate constant of 2 X 10(7) M-1 X S-1. Dissociation of the complex was characterized as a simple first-order process with rate constant on the order of 7 X 10(-3)S-1. The quotient of these is comparable to the dissociation constant as measured in equilibrium binding studies, 2.2 X 10(-10) M. The temperature dependence of the rate of association indicated an Arrhenius activation energy of 8.4 kcal X mol-1, while that of the equilibrium constant for dissociation indicated a standard enthalpy change of 8.8 kcal X mol-1. The large increase in affinity of nitrobenzylthioinosine as compared to natural nucleosides is attributable to an entropy-driven interaction with the binding site. Thymidine, dipyridamole and papaverine each decrease the apparent dissociation constant for the nitrobenzylthioinosine-cell complex; the latter, inhibitors of nucleoside transport, decrease the rate of dissociation of the complex.