Previous results [Tolkovsky, A. M., & Levitzki, A. (1978) in Hormones and Cell Regulation (Dumont, J., & Nunez, J., Eds.) Vol. 2, pp 89-105, Elsevier/North Holland Publishing Co., Amsterdam; (1978) Biochemistry 17, 3795-3810] indicated to us that the activation of turkey erythrocyte adenylate cyclase by the hormone-bound β-adrenergic receptor is a bimolecular process: [formula omitted] where HR is the hormone-bound receptor, E the inactive form of adenylate cyclase, E' the activated form of the enzyme, and k1 the bimolecular rate constant of enzyme receptor collision. (HRE) is the short-lived complex which does not accumulate. In this study we have designed experiments aimed to examine the possibility that this bimolecular process is diffusion controlled. Turkey erythrocyte membranes were fluidized progressively by the insertion of increasing amounts of cis-vaccenic acid. The fluidity of the native and the cis-vaccenic acid treated membrane was monitored by fluorescence polarization using diphenyl hexatriene as a fluorescence probe. In parallel, the rate constant of enzyme activation by the epinephrine-bound β receptor was measured. It was found that the bimolecular rate constant k1 increases linearly, 20-fold, as a function of membrane fluidity. This dependence of k1 on membrane fluidity strongly suggests that the process of adenylate cyclase activation by the β receptor is a diffusion controlled process. Applying the mathematical correlation between the bimolecular rate constant and the diffusion coefficient in two-dimensional space, one can calculate that the diffusion coefficient of the β-adrenergic receptor is in the range of 4.0X10-1 to 9.0X10-10 cm2/s at 25 °C. The increase in membrane fluidity also causes a maximal threefold increase of the adenylate cyclase specific activity. This effect is a direct effect on the catalytic unit, also found for the NaF-stimulated activity, and can be separated from the effect of membrane fluidity on the interaction between the receptor and the catalytic unit.
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