Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:4.6.1.2 (guanylate cyclase)
 8,497 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The subcellular localization of guanylate cyclase was examined in rat liver. About 80% of the enzyme activity of homogenates was found in the soluble fraction. Particulate guanylate cyclase was localized in plasma membranes and microsomes. Crude nuclear and microsomal fractions were applied to discontinuous sucrose gradients, and the resulting fractions were examined for guanylate cyclase, various enzyme markers of cell components, and electron microscopy. Purified plasma membrane fractions obtained from either preparation had the highest specific activity of guanylate cyclase, 30 to 80 pmol/min/mg of protein, and the recovery and relative specific activity of guanylate cyclase paralleled that of 5'-nucleotidase and adenylate cyclase in these fractions. Significant amounts of guanylate cyclase, adenylate cyclase, 5'-nucleotidase, and glucose-6-phosphatase were recovered in purified preparation of microsomes. We cannot exclude the presence of guanylate cyclase in other cell components such as Golgi. The electron microscopic studies of fractions supported the biochemical studies with enzyme markers. Soluble guanylate cyclase had typical Michaelis-Menten kinetics with respect to GTP and had an apparent Km for GTP of 35 muM. Ca-2+ stimulated the soluble activity in the presence of low concentrations of Mn-2+. The properties of guanylate cyclase in plasma membranes and microsomes were similar except that Ca-2+ inhibited the activity associated with plasma membranes and had no effect on that of microsomes. Both particulate enzymes were allosteric in nature; double reciprocal plots of velocity versus GTP were not linear, and Hill coefficients for preparations of plasma membranes and microsomes were calculated to be 1.60 and 1.58, respectively. The soluble and particulate enzymes were inhibited by ATP, and inhibition of the soluble enzyme was slightly greater. While Mg-2+ was less effective than Mn-2+ as a sole cation, all enzyme fractions were markedly stimulated with Mg-2+ in the presence of a low concentration of Mn-2+. Triton X-100 increased the activity of particulate fractions about 3- to 10-fold and increased the soluble activity 50 to 100%.
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PMID:Localization of particulate guanylate cyclase in plasma membranes and microsomes of rat liver. 23 12

Adenylate and guanylate cyclase activities were measured in rat small intestinal villus and crypt cells to determine possible correlations with cellular differentiation. Isolated intestinal cells were prepared by a method which effectively separates differentiated villus cells from undifferentiated crypt cells (J Biol Chem 248:2542, 1973). Crypt cells were found to have a significantly lower guanylate cyclase activity than villus cells. Adenylate cyclase activity was higher in crypt cells than villus cells, although the difference was less striking than the reverse gradient observed for guanylate cyclase. There was no gradient of activity for cyclic guanosine 3':5'-monophosphate phosphodiesterase. However, cyclic adenosine 3':5'-monophosphate phosphodiesterase activity was lower in villus cells. No villus to crypt gradient of cyclic adenosine 3':5'-monophosphate concentration was detected in mucosa frozen rapidly in liquid nitrogen. The properties and subcellular localization of the cyclases were also evaluated, and of particular interest was the localization of guanylate cyclase to the microvillus membrane and the confirmation of adenylate cyclase activity in the lateral-basal membrane. The villus to crypt gradient of guanylate cyclase suggests that this enzyme has a specialized role in the differentiated villus cell. The contrasting subcellular localization of the cyclases suggests that the cyclases may be interrelated, possibly reflecting the epithelial cell polarity for absorption and secretion.
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PMID:Adenylate and guanylate cyclase activities and cellular differentiation in rat small intestine. 23 99

Guanylate cyclase has been purified from extracts of Escherichia coli. After a 1000-fold purification, the enzyme contains only minor contaminants as judged by disc gel electrophoresis. The Km for GTP is approximately 7 times 10(-5) M and the optimal pH is 8.0. More activity is observed with Mn2+ than with Mg2+, and maximal activity is observed at 0.14 mM Mn2+ and 1.4 mM Mg2+. Based on its behavior on Sephadex G-100, the molecular weight of E. coli guanylate cyclase is about 30,000. Disc gel electrophoretic analysis indicates that the enzyme consists of a single polypeptide chain. Guanylate cyclase does not form 3':5'-AMP from ATP, and therefore, is distinct from adenylate cyclase.
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PMID:Guanylate cyclase in Escherichia coli. Purification and properties. 23 41

Guanylate cyclase from the rat renal medulla is found in both the soluble and particulate fractions of the cell. Sucrose density gradient centrifugation and gel filtration in H2O and D2O indicate that the enzyme from the soluble cell fraction has the following properties: S20w, 6.3 S; Stokes radius, 54 A; partial specific volume, 0.75 ml/g; mass, 154,000 daltons; f/fo, 1.4; axial ratio (prolate ellipsoid), 7. The addition of 0.1% Lubrol PX to this fraction activates the enzyme and changes thartial specific volume, 0.74 ml/g; mass, 148,000 daltons; f/fo, 1.6; axial ratio (prolate ellipsoid), 11. These findings show that detergent activates the enzyme by changing its conformation and not simply by dispersing nonsedimentable membrane fragments. The dimensions of this guanylate cyclase in detergent are very similar to those of detergent-solubilized adenylate cyclase from the same tissue (Neer, E.J. (1974) J. Biol. Chem. 249, 6527-6531). Guanylate cyclase can be solubilized from the particulate cell fraction with 1% Lubrol PX but has properties quite different from those of the guanylate cyclase in the soluble cell fraction. It is a large aggregate with a value of S20,w of about 10 S, Stokes radius of 65 A, and a mass of approximately 300,000 daltons. However, the peaks of guanylate cyclase activity in column effluents and sucrose density gradients are very broad indicating a mixture of different size proteins. The conditions used to solubilize guanylate cyclase from the particulate fraction also solubilize adenylate cyclase, and the two activities can be separated on the same sucrose gradient. Studies of this sort require a rapid, accurate guanylate cyclase assay. We have developed an assay for guanylate cyclase activity which meets these criteria by adapting the competitive protein binding assay for guanosine cyclic 3':5' monophosphate originally described by Murad et al. (Murad, F., Manganiello, V., and Vaughn, M. (1971) Proc. Natl. Acad. Sci. U.S.A. 68, 736-739).
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PMID:Guanylate cyclase from the rat renal medulla. Physical properties and comparison with adenylate cyclase. 24 Aug 41

At extremely low concentrations, in the picomole and the nanomole range, bradykinin produces contraction and relaxation of smooth muscle in the gastrointestinal and the urogenital tract. At the target organ, bradykinin interacts with discriminator proteins of the plasma membranes and triggers, via changes in certain membrane functions, its biological response:--The binding to the discriminator makes specific conformative and constitutional demands on the nonapeptide. The binding results from an angular conformation which exists in the solution. The complete sequence is responsible for this specific conformation. Consequently, the biological activity of partial sequences is low. The conformational analysis of analogues used in studies on the mechanism of action showed but slight differences from bradykinin. The interaction of these analogues with the discriminator protein is disturbed to a varying extent by modifications at positions 1, 5, 8 and 9 in the side chains. The affinity for the discriminator is affected, dependently on the respective configuration, by substitution on the beta-C atom in the two phenylalanine residues.--Bradykinin is not only bound to, but also degraded at, the plasma membranes of the rat uterus and duodenum. The bradykinin-degrading enzyme has been characterized as a kininase II with the aid of various inhibitors. The conformative and configurative prerequisites decisive for enzymatic degradation are others than those decisive for binding to the discriminator.--The changes in the activities of the membrane-bound adenylate and guanylate cyclases (produced by the bradykinin-discriminator complex) that take place at the rat duodenum and uterus in the presence of extracellular calcium ions contrast with each other: At the duodenum, the ratio between these two cyclic nucleotides is changed in favour of adenylate cyclase; and at the uterus, in favour of guanylate cyclase; Substances which increase or decrease the cAMP level may also potentiate or inhibit the relaxation of the duodenum. These bradykinin-induced changes in enzyme activity must be considered in connection with other effectors, e.g. prostaglandins and calcium ions.--The calcium-ion-dependence of the effect of bradykinin on the guinea-pig ileum and the rat uterus indicates the importance of these ions as additional second messengers. Bradykinin stimulates the influx of calcium ions into the ileum; it is ineffective if no extracellular calcium ions into the ileum; it is ineffective if no extracellular calcium ions are available. It seems that intracellular and membranal calcium is mobilized in the uterus, which is evidenced by results from experiments with EGTA on the isolated organ and by the release of calcium from plasma membranes after application of bradykinin. It is assumed that the observed changes in membrane functions are induced by the peptide-discriminator complex simultaneously and not in the form of a causal chain.
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PMID:[On the mode of action of bradykinin on smooth muscle (author's transl)]. 39 90

Current information on the intestinal secretory mechanism and on the actions of the enterotoxins from Vibrio cholerae and Escherichia coli are reviewed. These enterotoxins, through their effects on the metabolism of cyclic nucleotides, both inhibit active absorption and stimulate active secretion of water and electrolytes in the small intestine. the enterotoxin of V. chol erae and the heat-labile enterotoxin of E. coli affect these activities by stimulating adenylate cyclase, and the heat-stable enterotoxin of E. coli does so by stimulating guanylate cyclase.
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PMID:Modes of action of enterotoxins from Vibrio cholerae and EScherichia coli. 39 84

The size distribution of adenylate cyclase from the rat renal medulla solubilized with the nonionic detergents Triton X-100 and Lubrol PX was determined by gel filtration and by centrifugation in sucrose density gradients made up in H2O or D2O. The physical parameters of the predominant form in Triton X-100 are s20,w, 5.9S; Strokes radius, 62 A; partial specific volume (v), 0.74 ml/g; mass, 159,000 daltons; f/f0, 1.6; axial ratio (prolate ellipsoid), 11. For the minor form the values are: s20w, 3.0; Stokes radius, 28 A; mass, 38,000 daltons; f/f0, 1.2. The corresponding values determined in Lubrol PX are similar. The value for V for the enzyme indicates that it binds less than 0.2 mg detergent/mg protein. Since interactions with detergents probably substitute for interactions with lipids and hydrophobic amino acid side chains, these findings suggest that no more than 5% of the surface of adenylate cyclase is involved in hydrophobic interactions with other membrane components. Thus, most of the mass of the enzyme is not deeply embedded in the lipid bilayer of the plasma membrane. Similar studies have been performed on the soluble guanylate cyclase of the rat renal medulla. In the absence of detergent, the molecular properties of this enzyme are: s20w, 6.3S; Stokes radius, 54 A, V, 0.75 ml/g; mass, 154,000 daltons f/f0, 1.4; Axial ratio, 7. The addition of 0.1% Lubrol PX to this soluble enzyme increases it activity two- to fourfold and changes the physical properties to: s20,w, 5.5S; Stokes radius, 62 A; V, 0.74 ml/g; mass, 148,000 daltons, f/f0, 1.6; axial ratio, 11. These results show that Lubrol PX activates the enzyme by causing a conformational change with unfolding on the polypeptide chain. Guanylate cyclase from the particulate cell fraction can be solubilized with Lubrol PX but has properties quite different from those of the enzyme in the soluble cell fraction. It is a heterogeneous aggregate with s20,w, 10S; Stokes radius, 65 A; mass about 300,000 daltons. The conditions which solubilize guanylate cyclase also solubilize adenylate cyclase and the two activities can be separated on the same sucrose gradient.
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PMID:The size of adenylate cyclase and guanylate cyclase from the rat renal medulla. 125 62

The properties of brain capillary endothelial cells (BCECs) have been analyzed. BCECs express two types of receptor sites for endothelins (ETs), and ETA-like receptor, and an ETB-like receptor that is not coupled to phospholipase C but whose occupancy activates Na+/H+ exchange activity. The ETA receptor is positively coupled to phospholipase C and negatively coupled to adenylate cyclase. BCECs, unlike aortic endothelial cells, express high-affinity receptor sites for C-type natriuretic peptide. They respond to exogenous nitric oxide (NO) and to NO donor molecules by large activations of soluble guanylate cyclase. They produce little cGMP in response to A23187 or to agonists of phospholipase C but do so after an exposure to interleukin-1. The physiological consequence of the high reactivity of BCECs to vasoactive factors is discussed.
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PMID:Function of vasoactive factors in the cerebral microcirculation. 128 98

The squid giant axon has proved a useful model in the study of ionic channel gating, intracellular homeostasis and receptor-mediated signal transduction leading to generation of intracellular second messengers. In the latter category, previous studies on activation of adenylate or guanylate cyclase have used intact and intracellularly perfused axons to investigate the effects of extra- and intracellular agents on the transduction processes. However, the perfusion of the axon interior washes out many factors which may be important in the processes under study. We introduce here the use of porous cellulose dialysis tubing as a means to circumvent these problems. We find that this dialysis technique is a simple procedure to set-up, and the serotonin/G-protein/adenylate cyclase system can readily be studied in the dialysed axon. This approach should allow investigation under conditions which retain asymmetric transmembrane conditions.
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PMID:The use of intracellular dialysis to study signal transduction coupling in the squid giant axon. 132 34

Heat-stable enterotoxins activate guanylate cyclase, whereas heat-labile enterotoxins stimulate adenylate cyclase. Both classes of toxins cause secretory diarrhea at least in part by stimulating Cl- secretion in the intestine. The mechanism for regulation of Cl- secretion by guanosine 3',5'-cyclic monophosphate (cGMP) was investigated using cultured T84 intestinal cells as a model for intestinal crypt cells. Escherichia coli heat-stable enterotoxin (ST) markedly stimulated cGMP production in T84 cells. Cl- secretion across T84 cell monolayers cultured on permeable filters was stimulated by E. coli ST, cholera toxin, or 8-BrcAMP, but 8-BrcGMP was ineffective. cGMP analogues that are known to be potent and specific activators of cGMP-dependent protein kinase (cG-kinase) also had little effect on 36Cl- uptake by T84 cells cultured in plastic dishes. E. coli ST, forskolin, cholera toxin, or membrane-permeant cAMP analogues markedly increased 36Cl- uptake into T84 cells. The general protein kinase inhibitor, staurosporine, inhibited the stimulation of Cl- permeability elicited by E. coli ST, vasoactive intestinal peptide (VIP), or 8-BrcAMP. DEAE-Sephacel chromatography revealed a predominant type II isoform of cAMP-dependent protein kinase (cA-kinase) in T84 cells, whereas little or no cytosolic cG-kinase activity was found. Treatment of T84 cells with E. coli ST or VIP resulted in an increase in the cA-kinase activity ratio (-cAMP/+cAMP) if the cytosolic enzyme was assayed at reduced temperature (on ice).(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Stimulation of intestinal Cl- transport by heat-stable enterotoxin: activation of cAMP-dependent protein kinase by cGMP. 132 20


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