Adenosine interactions with thyroid adenylate cyclase

Wolff, J.; Londos, Constantine; Cook, G. Hope

doi:10.1016/0003-9861(78)90078-4

Cited by 44 publications

(6 citation statements)

References 31 publications

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“…In order to reduce the possibility that this implied effect at the level of the coupling mechanism was artifactual, we evaluated Gpp(NH)p sensitivity using two substrates: ribose ATP and deoxyribose ATP, shown to be an adenylate cyclase substrate more resistant to degradation to adenosine (Coopera Londos, 1979). Results obtained using this substrate are consequently free of ambiguities due to activation of the adenosine receptor and its commensurate guanyl nucleotide-dependent inhibitory effects on some hormone-sensitive adenylate cyclase systems (Wolff, Londos & Cook, 1978). Using deoxyribose ATP as substrate, the effect of PGE pretreatment on adenylate cyclase stimulation by Gpp(NH)p was still marked (Fig.…”

Section: Resultsmentioning

confidence: 95%

Interaction of prostaglandin E2 and β-adrenergic catecholamines in the regulation of uterine smooth muscle motility and adenylate cyclase in the rat

Krall¹,

Barrett²,

Jamgotchian³

et al. 1984

Journal of Endocrinology

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Prostaglandin E2 (PGE2) increased the force of the spontaneous contractions of the rat myometrium and decreased the sensitivity of the uterus to the relaxing effects of the specific beta-adrenergic catecholamine agonist isoprenaline. Prostaglandin E2, at concentrations above 10 mumol/l, increased cyclic AMP production by intact muscle strips. The muscle strips were far more sensitive, however, to the inhibitory effect PGE2 had on isoprenaline-dependent cyclic AMP production (threshold less than 0.001 nmol/l). Both PGE2 and isoprenaline stimulated adenylate cyclase activity of a myometrial subcellular (particulate) fraction in a guanyl nucleotide-requiring manner. When present in saturating concentrations (100 mumol/l), the stimulatory effects were not additive, suggesting that the receptors for the two agonists competed for the same catalytic subunit of adenylate cyclase or for the same guanyl nucleotide-requiring factor which couples receptors and enzyme. If muscle strips were incubated with PGE2 before the preparation of the adenylate cyclase-containing particulate fraction, the enzyme became less responsive to stimulation by guanyl nucleotide and by isoprenaline and PGE2 in the presence of guanyl nucleotide. The PGE2 receptor may therefore interact with the beta-adrenoreceptor to inhibit isoprenaline-dependent cyclic AMP production by intact muscle cells by desensitizing adenylate cyclase, possibly at the level of the guanyl nucleotide-dependent coupling step.

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Section: Resultsmentioning

confidence: 95%

Interaction of prostaglandin E2 and β-adrenergic catecholamines in the regulation of uterine smooth muscle motility and adenylate cyclase in the rat

Krall¹,

Barrett²,

Jamgotchian³

et al. 1984

Journal of Endocrinology

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“…Rat liver plasma membranes were prepared as described (25). Membranes from I-10 Ley-dig tumor cells, from Y-1 mouse adrenal tumors, and from beef thyroid were prepared as described (17,26). Human platelets were obtained from the National Institutes of Health Blood Bank; platelet membranes were prepared by freezing and thawing, as described by Haslam and Lynham (12).…”

Section: Methodsmentioning

confidence: 99%

Two distinct adenosine-sensitive sites on adenylate cyclase.

Londos¹,

Wolff²

1977

Proc. Natl. Acad. Sci. U.S.A.

485

228

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The effects of adenosine and adenosine analogs on adenylate cyclases from several tissues have been examined. Two adenosine-reactive sites have been identified: (i) the "R" site, occupancy of which usually leads to activation of cyclase and which requires integrity of the ribose ring for activity, and (ii) the "F" site, which mediates inhibition and requires integrity of the purine ring for activity. Biphasic effects of adenosine are explained by the presence of both sites on a single adenylate cyclase. Comparison of these data with those in the literature indicates that adenosine-reactive "P" and "R" sites are present generally.

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“…Although 2Ј-deoxyadenosine has an unfortunately high K i (240 Ϯ 60 M) compared with 2Ј-deoxy-3Ј-AMP (5 M, data not shown), we have utilized 2Ј-deoxyadenosine for this work because of the availability of the compound in radiolabeled form. Adenylyl cyclases (native enzymes or the soluble system utilized here) display noncompetitive or mixed noncompetitive inhibition by P-site analogs with respect to MnATP (3,4,10,11,15,23,24). We have focused our analysis on G s␣ -activated adenylyl cyclase activity with MgATP as substrate (in the absence of Mn 2ϩ ) to avoid the complications of mixed patterns of inhibition and the use of a second divalent cation (particularly Mn 2ϩ in the presence of PP i , which has limited solubility).…”

Section: P-site Inhibition Of Adenylylmentioning

confidence: 99%

The Catalytic Mechanism of Mammalian Adenylyl Cyclase

Dessauer

Gilman

1997

Journal of Biological Chemistry

122

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The mechanism of P-site inhibition of adenylyl cyclase has been probed by equilibrium binding measurements using 2-[ 3 H]deoxyadenosine, a P-site inhibitor, and by kinetic analysis of both the forward and reverse reactions (i.e. cyclic AMP and ATP synthesis, respectively). There is one binding site for 2-deoxyadenosine per C 1 /C 2 heterodimer; the K d is 40 ؎ 3 M. Binding is observed only in the presence of one of the products of the adenylyl cyclase reaction, pyrophosphate (PP i ). A substrate analog, Ap(CH 2 )pp (␣,␤-methylene adenosine 5-triphosphate), and cyclic AMP compete for the P-site in the presence of PP i , but P-site analogs do not compete for substrate binding (in the absence of PP i ). Kinetic analysis indicates that release of products from the enzyme is random. These facts permit formulation of a model for the adenylyl cyclase reaction, for which we provide substantial kinetic support. We propose that Psite analogs act as dead-end inhibitors of product release, stabilizing an enzyme-product (E-PP i ) complex by binding at the active site. Although product release is random, cyclic AMP dissociates from the enzyme preferentially. Release of PP i is slow and partially rate-limiting.Adenosine and various analogs of the nucleoside have both stimulatory and inhibitory effects on adenylyl cyclase activity (reviewed in Ref. 1). Londos and Wolff (2) categorized these effects mechanistically, based on their structure-activity relationships. Two types of adenosine-reactive sites were identified: those with strict requirements for the ribose moiety, designated R sites, and those with strict structural constraints for interaction with the purine ring, designated P sites. R sites are the ligand-binding sites of adenosine-specific G protein 1 -coupled receptors, which can either stimulate or inhibit adenylyl cyclase activity indirectly, while P sites, whose occupancy inhibits cyclic AMP synthesis, are structural features of adenylyl cyclases themselves (2-7). The physiological significance of Psite inhibition is unclear, but concentrations of 3Ј-AMP found in vivo appear sufficient to inhibit adenylyl cyclase activity (8).P-site inhibition is typically noncompetitive or uncompetitive with respect to substrate ATP, depending on the divalent cation utilized in the assay (Mn 2ϩ usually yielding noncompetitive kinetics; Mg 2ϩ uncompetitive) (3, 4, 9 -11). Furthermore, the apparent potency of such inhibitors increases when adenylyl cyclase is activated (3-5, 7, 11). Representative P-site reagents, ordered by potency, include 2Ј,5Ј-dideoxy-3Ј-ATP Ͼ 2Ј,5Ј-dideoxy-3Ј-ADP Ͼ 2Ј,5Ј-dideoxy-3Ј-AMP Ͼ 2Ј-deoxy-3Ј-AMP Ͼ 3Ј-AMP Ͼ 2Ј-deoxyadenosine Ͼ adenosine (12, 13). Although several mechanisms for P-site inhibition have been proposed, there is as yet no conclusive evidence to support any particular hypothesis. Using engineered, soluble forms of mammalian adenylyl cyclase, we and others have shown that the conserved cytosolic domains of the enzymes contain the structural components necessary for G s␣ -and forskolin-stimulate...

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Adenosine interactions with thyroid adenylate cyclase

Cited by 44 publications

References 31 publications

Interaction of prostaglandin E2 and β-adrenergic catecholamines in the regulation of uterine smooth muscle motility and adenylate cyclase in the rat

Interaction of prostaglandin E2 and β-adrenergic catecholamines in the regulation of uterine smooth muscle motility and adenylate cyclase in the rat

Two distinct adenosine-sensitive sites on adenylate cyclase.

The Catalytic Mechanism of Mammalian Adenylyl Cyclase

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