Fluoride stimulation of adenylate cyclase is dependent on the guanine nucleotide regulatory protein.

Adenylate cyclase in turkey erythrocyte membranes is activated only slowly by guanylyl-8,y-imidodiphosphate (GPP(NH)P) in the absence of isoproterenol but is responsive to fluoride. Incubation of membranes with isoproterenol and GMP allows subsequent isoproterenol-independent stimulation by GPP(NH)P and causes a decline in fluoride-stimulated enzyme activity. The decrement in fluoride activation is dependent upon time and temperature and requires both isoproterenol and GMP. The decline of fluoride-stimulated adenylate cyclase activity is reversed by subsequent incubation with GTP or by GDP or guanosine a,B-metbylene diphosphonate (GP(CH2)P). Fluoride does not prevent the hydrolysis of [3H]GTP to [%IGDP at isoproterenol-dependent regulatory sites, nor does fluoride enhance the conversion of [’HIGDP to [3H]GTP at such sites. An analog of GDP, guanosine 5’-0-(2-thiodiphosphate) (GDP-B-S) can compete with GPP(NJ3)P for occupancy of isoproterenol-dependent guanine nucleotide regulatory sites. Incubation of membranes with GDP-8-53 does not cause inhibition of fluoride-stimulated adenylate cyclase activity, but after incubation with GDP-/3-S and isoproterenol (enabling GDP-p-S to bind to the guanine nucleotide regulatory unit), fluoride stimulation of enzyme activity does not occur even if GTP is included in the incubation medium. We conclude that the guanine nucleotide regulatory protein is necessary for fluoride activation of adenylate cyclase and that the nature of the guanine nucleotide bound at the regulatory site influences fluoride stimulation. Exchange of the guanine nucleotide on the regulatory site with other nucleotides in the incubation medium is not necessary for fluoride stimulation of enzyme activity. Endogenous GDP, tightly bound to the guanine nucleotide regulatory protein, is sufficient for supporting fluoride stimulation of adenylate cyclase activity.