Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:4.6.1.1 (adenylate cyclase)
 19,190 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Controversy exists concerning the localization of the enzyme Na+,K(+)-ATPase to canalicular membranes in hepatocytes. Most studies find enzyme activity only at the basolateral plasma membrane domain of the hepatocyte. However, Na+,K(+)-ATPase activity has been detected recently in a canalicular membrane fraction prepared by Mg++ precipitation, suggesting that differences in membrane domain fluidity account for these discrepancies. To reinvestigate this question, we used free-flow electrophoresis to further purify canalicular liver plasma membranes originally separated by sucrose density centrifugation. With this technique, canalicular membranes devoid of Na+,K(+)-ATPase activity by routine assay were separated into six subfractions. More than 80% of the activities of canalicular marker enzymes was recovered in two subfractions closest to the anode, which were totally devoid of Na+,K(+)-ATPase activity. However, Na+,K(+)-ATPase activity could now be detected in the four other fractions that contained only small amounts of canalicular marker enzymes. The basolateral marker enzyme, glucagon-stimulated adenyl cyclase, comigrated with this cryptic Na+,K(+)-ATPase activity. Furthermore, addition of 6 mumol/L [12-(2-methoxyethoxy)-ethyl-8-(cis-2-n-octylcyclopropyl)-octanoate ], a membrane-fluidizing agent, to the original canalicular membrane preparation and to all subfractions did not stimulate or unmask latent Na+,K(+)-ATPase activity. Finally, when canalicular membranes isolated by Mg++ precipitation were subjected to free-flow electrophoresis, they could not be separated from the more positively charged Na+,K(+)-ATPase-containing fractions, probably because of alterations in surface charge. Together these findings suggest that Na+,K(+)-ATPase is a basolateral enzyme, that represents a small contaminant when present in canalicular liver plasma membranes and that methodological differences may account for previous discrepancies.
 ...
PMID:Cryptic Na+,K(+)-ATPase activity in rat liver canalicular plasma membranes: evidence for its basolateral origin. 215 68

Membrane proteins of transporting epithelia are often distributed between apical and basolateral surfaces to produce a functionally polarized cell. The distribution of Na+,K+-ATPase [ATP phosphohydrolase (Na+/K+-transporting), EC 3.6.1.37] between apical and basolateral membranes of hepatocytes has been controversial. Because Na+,K+-ATPase activity is fluidity dependent and the physiochemical properties of the apical membrane reduces its fluidity, we investigated whether altering membrane fluidity might uncover cryptic Na+,K+-ATPase in bile canalicular (apical) surface fractions free of detectable Na+,K+-ATPase and glucagon-stimulated adenylate cyclase activities. Apical fractions exhibited higher diphenylhexatriene-fluorescence polarization values when compared with sinusoidal (basolateral) membrane fractions. When 2-(2-methoxyethoxy)ethyl 8-(cis-2-n-octylcyclopropyl)octanoate (A2C) was added to each fraction, Na+,K+-ATPase, but not glucagon-stimulated adenylate cyclase activity, was activated in the apical fraction. In contrast, further activation of both enzymes was not seen in sinusoidal fractions. The A2C-induced increase in apical Na+,K+-ATPase approached 75% of the sinusoidal level. Parallel increases in apical Na+,K+-ATPase were produced by benzyl alcohol and Triton WR-1339. All three fluidizing agents decreased the order component of membrane fluidity. Na+,K+-ATPase activity in each subfraction was identically inhibited by the monoclonal antibody 9-A5, a specific inhibitor of this enzyme. These findings suggest that hepatic Na+,K+-ATPase is distributed in both surface membranes but functions more efficiently and, perhaps, specifically in the sinusoidal membranes because of their higher bulk lipid fluidity.
 ...
PMID:Biochemical localization of hepatic surface-membrane Na+,K+-ATPase activity depends on membrane lipid fluidity. 284 69

To assess whether internalization of beta-adrenergic receptor occurs in the CNS, we have isolated clathrin-coated vesicles from bovine forebrain and examined them for the presence of beta-adrenergic receptor binding and adenylate cyclase activities. A coated vesicle enriched preparation isolated by successive D2O-Ficoll density gradient centrifugations was applied to a glass bead permeation column to achieve further purification. Two major peaks of protein were eluted from the column and monitored by electron microscopy and SDS-PAGE. Peak II contained almost exclusively coated vesicles (98%), whereas peak I, which appeared in the void volume, contained larger smooth vesicles and few coated vesicles. beta-Adrenergic receptor binding to peaks I and II was measured with 125I-cyanopindolol (CYP) as ligand in Sepharose 4B column assays. 125I-CYP was found to bind specifically and saturably to both peaks I and II with a Bmax of 28 +/- 4 and 32 +/- 3 fmol/mg protein, respectively. 3H-CGP 12177, a hydrophilic beta-adrenergic receptor ligand, did not label receptors present in peak II, but it specifically bound to synaptic plasma membranes (SPM) prepared from bovine hippocampus and, to a lesser extent, to peak I. These results suggest that receptors present in coated vesicles are cryptic in nature. In the displacement of 125I-CYP binding by (-)-isoproterenol, addition of 50 microM GppNHp caused a significant "right shift" with SPM and peak I but not the peak II preparation. Adenylate cyclase activities could also be detected in both peaks I and II (specific activities, 21 +/- 0.6 and 24 +/- 0.5 pmol cAMP/mg protein/min, respectively).(ABSTRACT TRUNCATED AT 250 WORDS)
 ...
PMID:Detection and characterization of beta-adrenergic receptors and adenylate cyclase in coated vesicles isolated from bovine brain. 301 96

The conversion of phosphatidylethanolamine (PE) to phosphatidylcholine (PC) is catalyzed by two methyltransferases with S-adenosylmethionine as the methyl donor. PC formed by transmethylation is further metabolized by phospholipase A2. The synthesis and degradation of methylated phospholipids are involved in regulating the number of the beta-adrenergic receptors and their coupling to adenylate cyclase in rat reticulocytes, HeLa cells, and rat astrocytoma cells. Methylation of the phospholipids in these cells is stimulated by binding of agonists to the beta-adrenergic receptors. Accumulation of phosphatidyl-N-monomethylethanolamine causes an increase in membrane fluidity and enhances the coupling of the receptors to adenylate cyclase. Agents that inhibit phospholipid methylation decrease the number of receptors in intact HeLa cells, while increased phospholipid methylation unmasks cryptic receptors. Conversely, the degradation of methylated phospholipids appears to be closely associated with the desensitization of the beta-adrenergic receptors following prolonged stimulation with isoproterenol. Inhibition and stimulation of phospholipase A2 causes inhibition and stimulation of this desensitization process.
 ...
PMID:Phospholipid methylation: a possible mechanism of signal transduction across biomembranes. 627 27

The effects of two cryptic peptides from pro-TRH: Ps4 (160-169) and Ps5 (178-199) were investigated on basal and secretagogue (GRH and TRH)-induced releases of GH from perifused fragments of rat adenohypophysis. Validation of the perifusion system was done by measuring: (1) the dose-dependent effect of GRH and TRH on GH release; and (2) the stimulation of that release by forskolin (to mimic the adenylate cyclase pathway) or by phorbol ester (to mimic the protein kinase C pathway). We show that: (1) Ps4 and Ps5 (1 microM) do not modify basal GH release; (2) Ps4 (1 microM) changes neither GRH (10 nM)- nor TRH (100 nM)-induced release of GH; (3) Ps5 (100 nM and 1 microM) significantly decreases the release of GH induced by equimolar concentrations of TRH but not that induced by GRH.
 ...
PMID:A cryptic peptide of TRH prohormone inhibits TRH-induced GH release. 799 14

Inactivation of the gene encoding the periplasmic protease DegP confers a high-temperature-sensitive phenotype in Escherichia coli. We have previously demonstrated that a degP mutant of E. coli strain CBM (W3110 pldA1) is not temperature sensitive and showed that this was most likely due to constitutive activation of the sigma E and Cpx extracytoplasmic stress regulons in the parent strain. In this study, further characterization of this strain revealed a previously unknown cryptic mutation that rescued the degP temperature-sensitive phenotype by inducing the extracytoplasmic stress regulons. We identified the cryptic mutation as an 11-bp deletion of nucleotides 1884 to 1894 of the adenylate cyclase-encoding cyaA gene (cyaAdelta11). The mechanism in which cyaAdelta11 induces the sigma E and Cpx regulons involves decreased activity of the mutant adenylate cyclase. Addition of exogenous cyclic AMP (cAMP) to the growth medium of a cyaAdelta11 mutant strain that contains a Cpx- and sigma E-inducible degP-lacZ reporter fusion decreased beta-galactosidase expression to levels observed in a cyaA+ strain. We also found that a cyaA null mutant displayed even higher levels of extracytoplasmic stress regulon activation compared to a cyaAdelta11 mutant. Thus, we conclude that the lowered concentration of cAMP in cyaA mutants induces both sigma E and Cpx extracytoplasmic stress regulons and thereby rescues the degP temperature-sensitive phenotype.
 ...
PMID:Adenylate cyclase mutations rescue the degP temperature-sensitive phenotype and induce the sigma E and Cpx extracytoplasmic stress regulons in Escherichia coli. 1615 63

Pseudomonas syringae pv. tomato DC3000 is a pathogen of tomato and Arabidopsis that translocates virulence effector proteins into host cells via a type III secretion system (T3SS). Many effector-encoding hypersensitive response and pathogenicity (Hrp) outer protein (hop) genes have been identified previously in DC3000 using bioinformatic methods based on Hrp promoter sequences and characteristic N-terminal amino acid patterns that are associated with T3SS substrates. To approach completion of the Hop/effector inventory in DC3000, 44 additional candidates were tested by the Bordetella pertussis calmodulin-dependent adenylate cyclase (Cya) translocation reporter assay; 10 of the high-probability candidates were confirmed as T3SS substrates. Several previously predicted hop genes were tested for their ability to be expressed in an HrpL-dependent manner in culture or to be expressed in planta. The data indicate that DC3000 harbors 53 hop/avr genes and pseudogenes (encoding both injected effectors and T3SS substrates that probably are released to the apoplast); 33 of these genes are likely functional in DC3000, 12 are nonfunctional members of valid Hop families, and 8 are less certain regarding their production at functional levels. Growth of DC3000 in tomato and Arabidopsis Col-0 was not impaired by constitutive expression of repaired versions of two hops that were disrupted naturally by transposable elements or of hop genes that are naturally cryptic. In summary, DC3000 carries a complex mixture of active and inactive hop genes, and the hop genes in P. syringae can be identified efficiently by bioinformatic methods; however, a precise inventory of the subset of Hops that are important in pathogenesis awaits more knowledge based on mutant phenotypes and functions within plants.
 ...
PMID:Multiple approaches to a complete inventory of Pseudomonas syringae pv. tomato DC3000 type III secretion system effector proteins. 1707 1