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Cyclase

About: Cyclase is a research topic. Over the lifetime, 10162 publications have been published within this topic receiving 388566 citations.


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Journal ArticleDOI
TL;DR: Results demonstrate that both fragments of the 43-kDa form of adenylate cyclase are essential for a high level of enzymatic activity.

99 citations

Journal Article
TL;DR: This work delineated in detail the binding specificity of the adenylate cyclase-coupled beta adrenergic receptors in a model system, the frog erythrocyte membrane, and found that agonists and antagonists appeared to compete for the same set of receptor binding sites.
Abstract: Recently developed techniques for directly studying ligand binding to beta adrenergic receptors with (-)-[3H]alprenolol have been used to delineate in detail the binding specificity of the adenylate cyclase-coupled beta adrenergic receptors in a model system, the frog erythrocyte membrane. The abilities of 60 beta adrenergic agents to compete for the binding sites and to interact with the adenylate cyclase (as agonists or antagonists) were quantitated and compared. The specificity of the receptors determined by direct binding studies or by adenylate cyclase studies was comparable. The KD values of the agents as determined by inhibition of (-)-[3H]alprenolol binding correlated well ( r = 0.95) with their apparent dissociation constants determined by enzyme studies. The latter were determined as the concentrations of agonists necessary to cause 50% maximal enzyme stimulation, or the concentrations of antagonists necessary to produce a 2-fold rightward shift in the (-)-isoproterenol dose-response curve. Agonists and antagonists appeared to compete for the same set of receptor binding sites. Structure-activity relationships determined by the direct binding studies were in excellent agreement with those previously determined in more intact tissue preparations. For agonists the structural features which determined receptor affinity (assessed by direct binding studies) were distinct from those which determined intrinsic activity (maximum ability to stimulate adenylate cyclase). The affinity of agonists was increased by increasing the size of the substituent on the amino nitrogen, by a (-) configuration of the hydroxyl on the β-carbon, and by the presence of a catechol moiety. Methyl or ethyl substitution on the α-carbon had only a slight (generally inhibitory) effect on affinity. Intrinsic activity of agonists was determined primarily by the nature of the substituents on the phenyl ring. Full intrinsic activity requires the presence of hydroxyl groups on the ring at positions 3 and 4 as well as the β-carbon hydroxyl in the (-) configuration. Deletion of the β-carbon hydroxyl, as in compounds such as dopamine, dobutamine, and related agents, leads to substantial loss of intrinsic activity and affinity even in the presence of large amino nitrogen substituents. A methanesulfonamide group substituted for the hydroxyl in position 3 on the ring results in reduced intrinsic activity. Deletion of the ring hydroxyl at either position 3 or 4 or substitution by chlorine produces competitive antagonists. Structure-activity relationships of antagonists were similar to those of agonists, except that the catechol moiety was replaced by a single or double aromatic ring structure. Separation of this moiety from the ethanolamine side chain by an ether function significantly increased affinity. When a phenyl group was present, a single substituent at the para position was associated with reduced affinity. ACKNOWLEDGMENT We gratefully acknowledge the expert assistance of the analytical chemistry department of New England Nuclear Corporation in developing chromatographic procedures for (-)-alprenolol.

99 citations

Journal ArticleDOI
TL;DR: These results provide the first direct evidence that a mutation linked to congenital blindness increases Ca2+ in the outer segment, which may trigger the apoptotic process.
Abstract: Guanylyl cyclase-activating proteins (GCAPs) are Ca2+-binding proteins that activate guanylyl cyclase when free Ca2+ concentrations in retinal rods and cones fall after illumination and inhibit the cyclase when free Ca2+ reaches its resting level in the dark. Several forms of retinal dystrophy are caused by mutations in GUCA1A, the gene coding for GCAP1. To investigate the cellular mechanisms affected by the diseased state, we created transgenic mice that express GCAP1 with a Tyr99Cys substitution (Y99C GCAP1) found in human patients with a late-onset retinal dystrophy ([Payne et al., 1998][1]). Y99C GCAP1 shifted the Ca2+ sensitivity of the guanylyl cyclase in photoreceptors, keeping it partially active at 250 nm free Ca2+, the normal resting Ca2+ concentration in darkness. The enhanced activity of the cyclase in the dark increased cyclic nucleotide-gated channel activity and elevated the rod outer segment Ca2+ concentration in darkness, measured by using fluo-5F and laser spot microscopy. In different lines of transgenic mice the magnitude of this effect rose with the Y99C GCAP1 expression. Surprisingly, there was little change in the rod photoresponse, indicating that dynamic Ca2+-dependent regulation of cGMP synthesis was preserved. However, the photoreceptors in these mice degenerated, and the rate of the cell loss increased with the level of the transgene expression, unlike in transgenic mice that overexpressed normal GCAP1. These results provide the first direct evidence that a mutation linked to congenital blindness increases Ca2+ in the outer segment, which may trigger the apoptotic process. [1]: #ref-36

99 citations

Journal ArticleDOI
TL;DR: Findings indicate that the B96Bom receptor is a B. mori TA receptor, which is negatively coupled to adenylate cyclase, and the use of this expression system should facilitate physiological studies of TA receptors as well as structure–activity studies ofTA receptor ligands.
Abstract: A cDNA encoding a biogenic amine receptor (B96Bom) was isolated from silkworm (Bombyx mori) larvae, and the ligand response of the receptor stably expressed in HEK-293 cells was examined. Tyramine (TA) at 0.1-100 micro m reduced forskolin (10 micro m)-stimulated intracellular cAMP levels by approximately 40%. The inhibitory effect of TA at 1 micro m was abolished by yohimbine and chlorpromazine (each 10 micro m). Although octopamine (OA) also reduced the cAMP levels, the potency was at least two orders of magnitude lower than that of TA. Furthermore, unlabelled TA (IC50 = 5.2 nm) inhibited specific [3H]TA binding to the membranes of B96Bom-transfected HEK-293 cells more potently than did OA (IC50 = 1.4 micro m) and dopamine (IC50 = 1.7 micro m). Taken together with the result of phylogenetic analysis, these findings indicate that the B96Bom receptor is a B. mori TA receptor, which is negatively coupled to adenylate cyclase. The use of this expression system should facilitate physiological studies of TA receptors as well as structure-activity studies of TA receptor ligands.

99 citations

Journal ArticleDOI
TL;DR: Although farnesol induces the accumulation of intracellular reactive oxygen species (ROS), ROS generation is not necessary for the induction of catalase (Cat1)-mediated oxidative-stress resistance.
Abstract: Farnesol, a Candida albicans cell-cell signaling molecule that participates in the control of morphology, has an additional role in protection of the fungus against oxidative stress. In this report, we show that although farnesol induces the accumulation of intracellular reactive oxygen species (ROS), ROS generation is not necessary for the induction of catalase (Cat1)-mediated oxidative-stress resistance. Two antioxidants, α-tocopherol and, to a lesser extent, ascorbic acid effectively reduced intracellular ROS generation by farnesol but did not alter farnesol-induced oxidative-stress resistance. Farnesol inhibits the Ras1-adenylate cyclase (Cyr1) signaling pathway to achieve its effects on morphology under hypha-inducing conditions, and we demonstrate that farnesol induces oxidative-stress resistance by a similar mechanism. Strains lacking either Ras1 or Cyr1 no longer exhibited increased protection against hydrogen peroxide upon preincubation with farnesol. While we also observed the previously reported increase in the phosphorylation level of Hog1, a known regulator of oxidative-stress resistance, in the presence of farnesol, the hog1/hog1 mutant did not differ from wild-type strains in terms of farnesol-induced oxidative-stress resistance. Analysis of Hog1 levels and its phosphorylation states in different mutant backgrounds indicated that mutation of the components of the Ras1-adenylate cyclase pathway was sufficient to cause an increase of Hog1 phosphorylation even in the absence of farnesol or other exogenous sources of oxidative stress. This finding indicates the presence of unknown links between these signaling pathways. Our results suggest that farnesol effects on the Ras-adenylate cyclase cascade are responsible for many of the observed activities of this fungal signaling molecule.

98 citations


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Performance
Metrics
No. of papers in the topic in previous years
YearPapers
202324
202257
202145
202048
201939
201856