Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.4.1 (phosphodiesterase)
 18,767 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

PC-1 is an ecto-enzyme possessing alkaline phosphodiesterase I (EC 3.1.4.1) and nucleotide pyrophosphatase (EC 3.6.1.9) activities. It has also been proposed to be an ecto-protein kinase capable of phosphorylating itself as well as exogenous proteins. We have investigated the phosphorylation capability of PC-1 and have developed a novel method for its detection and characterization based on autophosphorylation, which allows detection without the use of antibodies. When cells expressing membrane PC-1 were held on ice with [gamma-32P]ATP, SDS/PAGE of whole cell lysates showed a single band which was PC-1; this band was absent in cells not expressing PC-1. Immunoprecipitates of soluble PC-1 isolated from culture supernatants of cells expressing PC-1 were also capable of autophosphorylation, and the size of the labeled protein was the same as previously reported for soluble PC-1. PC-1 was also labeled with [alpha-32P]ATP and [35S]dATP[alpha S]. We found no evidence that PC-1 was capable of phosphorylating proteins other than itself, and conclude that it is not a true kinase, and that the observed labeling with [gamma-32P]ATP, [alpha-32P]ATP and [35S]dATP[alpha S] reflect transient covalent adducts that are part of the catalytic cycle of phosphodiesterase/pyrophosphatase activity rather than intrinsic kinase activity. Mutation of the active-site threonine to tyrosine, serine or alanine reduced the 5'-nucleotide phosphodiesterase activity of PC-1 and its ability to autophosphorylate to undetectable levels. Together, these data suggest that both activities depend on the same site.
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PMID:Autophosphorylation of PC-1 (alkaline phosphodiesterase I/nucleotide pyrophosphatase) and analysis of the active site. 773 62

The plasma cell-membrane glycoprotein PC-1 is an ectoenzyme with alkaline phosphodiesterase I/5'-nucleotide phosphodiesterase (EC 3.1.4.1) and nucleotide pyrophosphatase (EC 3.6.1.9) activities. It contains sequence motifs which closely match the consensus EF-hand (helix-loop-helix) Ca(2+)-binding regions of parvalbumin, troponin-C and calmodulin, and its enzymic activity is increased in the presence of divalent cations and decreased in the presence of chelating agents. We have undertaken experiments to determine whether divalent cations affect the conformation of the PC-1 protein, as assessed by their effect on thermal stability, resistance to proteolysis and binding of polyclonal antibodies to the whole native protein and monoclonal antibodies to a putative Ca(2+)-binding region. Divalent cations were found to protect solubilized PC-1 against thermal denaturation and proteolysis. They also stabilized PC-1 on intact cells; this form was much more resistant to proteolysis than Triton X-100 solubilized PC-1. Ca2+, Mg2+ and Zn2+ ions were equally effective. Monoclonal antibodies to the bacterially expressed C-terminal EF-hand homology region only bound to mammalian PC-1 in the absence of Ca2+. In contrast, the great majority of polyclonal antibodies to native PC-1 bound regardless of whether Ca2+ was present or not, but with increased binding when Ca2+ was present. These results provide evidence that divalent cations bind to PC-1 and stabilize its conformation.
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PMID:Divalent cations stabilize the conformation of plasma cell membrane glycoprotein PC-1 (alkaline phosphodiesterase I). 799 60

Recent studies have suggested that the insulin receptor tyrosine kinase inhibitor, membrane glycoprotein PC-1, may play a role in certain insulin resistant states. In the present study, we examined whether either insulin receptor function or PC-1 activity was altered during the development of insulin resistance that occurs with high fat feeding in normal rats. Over the course of 14 days of high fat feeding, both maximal and submaximal (physiological) insulin-stimulated skeletal muscle glucose uptake decreased gradually; after 14 days of high fat feeding, submaximal and maximal insulin-stimulated glucose uptake decreased by approximately 40 and approximately 50%, respectively. In contrast, in the same muscles (tibialis anterior) of these animals, neither insulin receptor content nor insulin-stimulated insulin receptor autophosphorylation was altered after 14 days of high fat feeding. PC-1 has both nucleotide pyrophosphatase (EC 3.6.1.9) and alkaline phosphodiesterase I (EC 3.1.4.1) enzyme activities. These enzyme activities showed no changes during the course of 14 days of high fat feeding. Individual data revealed that there was no significant correlation between insulin-stimulated glucose uptake and alkaline phosphodiesterase or nucleotide pyrophosphatase activity (P > 0.05). Together, these data indicate that neither defects in insulin receptor function nor elevated PC-1 activities are involved in the development of insulin resistance in rats with high fat feeding, and the insulin resistance induced with high fat feeding is likely due to postreceptor defects in skeletal muscle.
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PMID:The development of insulin resistance with high fat feeding in rats does not involve either decreased insulin receptor tyrosine kinase activity or membrane glycoprotein PC-1. 898 41

NAD may be degraded in several ways. A large number of investigations have shown that at least those catabolic routes which involve the formation of ADP-ribose are related to regulatory processes. In this study a rapid assay was utilized that permits identification of NAD-degrading enzymes directly in sodium dodecylsulfate polyacrylamide gels. Enzymatic activities were recovered by washing the gels in the presence of mild detergents such as lauryl dimethylamine N-oxide or Triton X-100. Subsequent incubation of the gels in the presence of the fluorescent analog 1,N6 etheno-NAD visualized NAD-degrading enzymes. Following excision of the fluorescent bands from the gels, the actual activity of the proteins was established by incubating the gel slices with 14C-labeled NAD and subsequent product analysis by thin layer chromatography (TLC). Homogenates from rat renal cortex and spleen were analyzed by this procedure. While in the spleen homogenate only a single band could be 'activity-stained', in the kidney three bands were detected. Kidney proteins with apparent molecular masses of about 210,000 and 105,000 Da were identified as phosphodiesterase and NAD pyrophosphatase (alkaline phosphodiesterase I), respectively. The third protein exhibited an apparent molecular mass of 41,000. The spleen protein (apparent molecular mass 45,000 Da) cleaved NAD to nicotinamide and ADP-ribose identifying it as NAD glycohydrolase. The procedure is suitable to screen for NAD-converting activities in crude extracts. It is specific for proteins which function as monomers or homo-oligomers.
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PMID:Detection and identification of NAD-catabolizing activities in rat tissue homogenates. 921 9

Vascular smooth muscle cell hypertrophy and proliferation may participate in the pathophysiology of cardiovascular disease. The analysis of changes in gene expression in vascular smooth muscle cells is crucial to the understanding of the molecular biology of cardiovascular disease. An effective method for analysis of gene expression is the differential display approach. Applying the differential display approach, we identified a gp130RB13-6-related gene in vascular smooth muscle cells following stimulation with platelet-derived growth factor-BB and angiotensin II. It is well known that gp130RB13-6 is a phosphodiesterase/nucleotide pyrophosphatase. Northern blotting and reverse transcriptase-polymerase chain reaction analysis revealed a dramatic down-regulation of the gp130RB13-6-related mRNA after six hours of stimulation of the cells with both agonists. Recently, gp130RB13-6 was identified as a rat neural differentiation and tumor cell surface plasma membrane glycoprotein. These findings demonstrate that the expression of gp130RB13-6 mRNA in vascular smooth muscle cells is remarkably regulated by growth factors and therefore may play an important role in the regulation of vascular smooth muscle cell growth.
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PMID:Identification of a phosphodiesterase I/nucleotide pyrophosphatase-related gene mRNA in rat vascular smooth muscle cells by the differential display approach. 964 40

Because dinucleotides are signaling molecules that can interact with cell surface receptors and regulate the rate of mucociliary clearance in lungs, we studied their metabolism by using human airway epithelial cells. A membrane-bound enzyme was detected on the mucosal surface of polarized epithelia that metabolized dinucleotides with a broad substrate specificity (diadenosine polyphosphates and diuridine polyphosphates [Up(n)U], n = 2 to 6). The enzymatic reaction yielded nucleoside monophosphates (NMP) and Np(n)(-)(1) (N = A or U), and was inhibited by nucleoside 5'-triphosphates (alpha,betamet adenosine triphosphate [ATP] > ATP >/= uridine triphosphate > guanidine triphosphate > cytidine triphosphate). The apparent Michaelis constant (K(m,app)) and apparent maximal velocity (V(max,app)) for [(3)H]Up(4)U were 22 +/- 4 microM and 0.24 +/- 0.05 nmoles. min(-)(1). cm(-)(2), respectively. Thymidine 5'-monophosphate p-nitrophenyl ester and adenosine diphosphate (ADP)- ribose, substrates of ecto alkaline phosphodiesterase I (PDE I) activities, were also hydrolyzed by the apical surface of airway epithelia. ADP-ribose competed with [(3)H]Up(4)U, with a K(i) of 23 +/- 3 microM. The metabolism of ADP-ribose and Ap(4)A was not affected by inhibitors of cyclic nucleotide phosphodiesterases (3-isobutyl-1-methylxanthine, Ro 20-1724, and 1,3-dipropyl-8-p-sulfophenylxanthine), but similarly inhibited by fluoride and N-ethylmaleimide. These results suggest that a PDE I is responsible for the hydrolysis of extracellular dinucleotides in human airways. The wide substrate specificity of PDE I suggests that it may be involved in several signaling events on the luminal surface of airway epithelia, including purinoceptor activation and cell surface protein ribosylation.
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PMID:Biochemical evidence for an ecto alkaline phosphodiesterase I in human airways. 1091 94


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