Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.2.1.17 (lysozyme)
21,489 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We studied the effect of adenosine nucleotides on several aspects of the functional activation of human peripheral blood polymorphonuclear leukocytes (PMN). Radiolabeled ATP bound to PMN in a manner suggesting the existence of specific binding sites because: 1) binding was reversed (92 +/- 6%) by 100-fold excess concentrations of unlabeled ATP but minimally by either ADP (43 +/- 12%) or GTP (37 +/- 8%); and 2) binding saturation was achieved (i.e., specific binding did not increase) above 250 microM ATP. Binding studies revealed that significant ATP hydrolysis occurred, even at low temperatures and in the presence of phosphatase inhibitors. Adenosine nucleotides activated signal transduction mechanisms in PMN because: 1) 1 to 100 microM ATP and 5'-adenylylimidodiphosphate (AMP-PNP) stimulated increased production of 1,2-diacylglycerols; 2) ATP (0.5 to 500 microM) and ADP (0.1 to 10 mM) induced increased insoluble protein kinase (PKC) activity in a dose-dependent manner when used at concentrations greater than 50 microM; 3) ATP (greater than or equal to 50 microM) induced a shift in the solubility of phorbol receptors from mostly soluble (89% in untreated cells) to mostly insoluble (68%), whereas ADP, GTP, and GDP were effective at higher concentrations; and 4) greater than or equal to 50 microM ATP stimulated increased phosphorylation of endogenous PMN proteins. AMP-PNP induced PKC activity and phosphoprotein changes that were qualitatively similar to those observed when PMN were treated with ATP, suggesting that extracellular ATP hydrolysis is not required for signal transduction to activate PKC. Functionally, ATP stimulated the secretion of specific (but not azurophil) granules because vitamin B12-binding protein and low levels of lysozyme, but not beta-glucuronidase, were released; qualitatively similar results were obtained by using AMP-PNP. These results suggest that certain adenosine nucleotides employed at physiologically relevant concentrations stimulate increased 1,2-diacylglycerol production, PKC activity, granule secretion, and endogenous phosphoprotein formation in a manner that is independent of extracellular ATP hydrolysis.
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PMID:Extracellular adenosine nucleotides stimulate protein kinase C activity and human neutrophil activation. 215 72

Two gastrin analogs containing a D- and a L-tetrafluorinated tyrosyl residue (Arg-Arg-Leu-Glu-Glu-Glu-Glu-Glu-Ala-(F4)Tyr-Gly) were synthesized and tested as substrates and inhibitors of the insulin receptor kinase. No phosphorylation of these peptides was observed, but both gastrin analogs were effective inhibitors in the microM range. Although the D- and L-tetrafluorotyrosine-gastrin analogs differ in the sequence by only 1 amino acid residue, a different inhibitory pattern was obtained with the insulin receptor. The inhibition of all-L-isomer is competitive with respect to both the protein substrate, reduced, S-carboxymethylated, and maleylated lysozyme (RCMM-lysozyme), and ATP with a Ki value of 4 microM. This result corroborates a previous finding (Walker, D. H., Kuppuswamy, D., Visvanathan, A., and Pike, L. J. (1987) Biochemistry 26, 1428-1433) that the kinetic mechanism for insulin receptor is a random Bi Bi mechanism. Different from the L-isomer, the D-analog is competitive to RCMM-lysozyme and noncompetitive toward ATP and gives an apparent inhibition constant of 20 microM. A free tetrafluorotyrosine also shows a competitive inhibition to protein substrate, RCMM-lysozyme (Ki = 18 mM) whereas free tyrosine shows no effect on the activity of insulin receptor. These results show the importance of the charge state and nucleophilicity of the phenolic component in substrate recognition and catalysis and provide a rationale for the design of inhibitors of tyrosyl phosphorylation.
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PMID:A rationale for the design of an inhibitor of tyrosyl kinase. 216 84

Mammalian cells contain two large proteolytic complexes, the 650-kDa proteasome (or multicatalytic protease) and the 1500-kDa (26 S) Ubiquitin-conjugate-degrading enzyme. Since the proteasome is also required for the ATP-dependent degradation of ubiquitinated proteins, we tested whether it may be a component of the larger complex. The proteasome normally is soluble in 38% ammonium sulfate. However, after preincubation of reticulocyte extracts with ATP, several proteasome activities appeared in the 38% ammonium sulfate pellet, including the ability to degrade hydrophobic peptides and 14C-casein. Also, following preincubation with ATP, the precipitable fraction could degrade 125I-lysozyme-ubiquitin (Ub) conjugates. The activities were not present after incubation without ATP or with a nonmetabolizable ATP analog. Nondenaturing gel electrophoresis indicated the ATP-dependent appearance of a new band which degraded proteasome substrates, and reacted with an anti-proteasome monoclonal antibody on Western blot. This new band appeared larger than the proteasome and migrated similarly to the larger Ub-conjugate-degrading complex. The formation of the larger complex required factor(s) present in the 38% ammonium sulfate pellet and either the 40-80% fraction or the purified proteasome from reticulocytes or muscle. After complex formation, hydrolysis of Ub-protein conjugates and also the non-ubiquitinated substrate, casein, was stimulated severalfold by ATP, but non-metabolizable ATP analogs had little or no effect. Thus, the proteasome corresponds to component CF-3 of Ganoth et al. (Ganoth, D., Leshinisky, E., Eytan, E., and Hershkov, A. (1989) J. Biol. Chem. 263 12412-12419) and undergoes an energy-dependent association with other factors to form the 1500-kDa, ATP-requiring proteolytic complex.
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PMID:The proteasome (multicatalytic protease) is a component of the 1500-kDa proteolytic complex which degrades ubiquitin-conjugated proteins. 218 Sep 50

A full-length Caenorhabditis elegans cDNA that encodes the alpha subunit of casein kinase II was inserted into the inducible bacterial expression vector pET3a to generate the plasmid pCK alpha. Escherichia coli DE21 lysozyme S that was transformed with pCK alpha expressed soluble, catalytically active casein kinase II alpha upon induction with isopropyl beta-D-thiogalactopyranoside. The expressed alpha subunit was purified to homogeneity with a 60% yield by chromatography on CM-Sephadex, P-11 phosphocellulose, and heparin-agarose. The Mr values estimated from sodium dodecyl sulfate-polyacrylamide gel electrophoresis (Mr = 42,000) or calculated from hydrodynamic measurements (s20,w = 3.3 S, Stokes radius = 2.8 nm, Mr = 37,000) were similar, thereby indicating that the expressed enzyme is monomeric. The native holoenzyme and the expressed alpha subunit exhibited several similar properties including the utilization of both ATP and GTP as substrates and the susceptibility to inhibition of phosphotransferase activity by low concentrations of heparin. However, the kcat for E. coli-derived alpha was only 9% of the kcat for the native holoenzyme, and catalytic activity was not stimulated by polyamines. Recombinant casein kinase II alpha aggregates at low ionic strength, and the aggregation is partially reversible. A mutant alpha subunit in which Lys74 and Lys75 were substituted by glutamic acid residues was constructed by site-directed mutagenesis. The mutant enzyme was not inhibited by typically effective concentrations of heparin (e.g. IC50 = 0.3 micrograms/ml) because the affinity of modified recombinant casein kinase II Glu-74Glu-75 for heparin decreased approximately 70-fold. Thus, Lys74 and Lys75 are implicated in the heparin binding, inhibitory domain. The successful expression of casein kinase II alpha in E. coli will facilitate the analysis of the structural basis for functional domains in this enzyme.
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PMID:Expression of wild-type and mutated forms of the catalytic (alpha) subunit of Caenorhabditis elegans casein kinase II in Escherichia coli. 224 6

Human oesophageal submucosal glands may be regularly demonstrated by first exposing the oesophageal lumen to toluidine blue which reveals the duct ostia. Four types of cell were identified in the glands - mucous, subsidiary or serous, myoepithelial and oncocytes. The mucous cell contained neutral, sialated and sulphated mucins. The subsidiary cells held smaller amounts of neutral and sialated mucin, plus fucosyl residues. No lipids were detectable histochemically. ATP-ase and alkaline phosphatase were shown in the capillary endothelium. The duct epithelium showed some nonspecific esterase activity not sensitive to E 600. By immunoperoxidase techniques, the duct epithelium was shown to be rich in cytokeratin. The subsidiary cells contained lysozyme, CEA and pepsinogen. B lymphocytes composed most of the periductular lymphoid aggregates, although some T cells were found there and also intraepithelial and subepithelial in relation to the stratified squamous epithelium lining the oesophagus. Langerhans' cells were also demonstrated as intraepithelial by several techniques.
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PMID:Human oesophageal submucosal glands. Their detection mucin, enzyme and secretory protein content. 243 35

Treatment of rabbit polymorphonuclear leukocytes (PMNs) with 20 mM sodium fluoride for 10 min, followed by removal of fluoride and addition of Ca2+ results in extensive exocytosis. This is apparent from a strong lysozyme release, together with a slight LDH release. During fluoride-activated Ca2+-dependent exocytosis an increase of indo fluorescence and a strong association of 45Ca with the cells occurs. Different inhibitors inhibit both 45Ca association and lysozyme release. Pretreatment of PMNs with pertussis toxin, or the presence of AI3+ in the medium has little effect on fluoride-activated Ca2+-dependent exocytosis. During pretreatment with fluoride, the ATP level strongly decreases. Exocytosis nevertheless occurs upon addition of Ca2+, indicating that a normal ATP level is not required for exocytosis. The glycogen content of the cell strongly decreases during exposure to Ca2+ after pretreatment with fluoride, but not during pretreatment with fluoride. Breakdown of glycogen and accumulation of 3-phosphoglycerate suggest that glycolysis is blocked at the enolase step, but proceeds as far as that step.
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PMID:Exocytotic enzyme release from rabbit polymorphonuclear leukocytes after treatment with fluoride and calcium. 250 51

Eukaryotic cells contain a 700-kDa proteolytic complex (the "proteasome" or multicatalytic endopeptidase complex), whose role in intracellular protein breakdown is unclear. It has been suggested that the proteasome functions in the rapid degradation of oxidant-damaged proteins and in the ATP-dependent proteolytic pathway. To test these possibilities, oxidant-damaged hemoglobin and albumin were produced by treating hemoglobin and albumin with phenylhydrazine, with hydroxyl radicals, or with both hydroxyl and superoxide radicals. After oxidant damage, these proteins were degraded more rapidly in erythrocyte extracts and also by the purified proteasome. However, complete removal of proteasomes from these extracts by immunoprecipitation (or inhibitors of its proteolytic activity) did not reduce the breakdown of oxidant-damaged hemoglobin and decreased degradation of hydroxyl- and superoxide-treated proteins by only 30-40%. Thus, erythrocytes must contain another proteolytic system for degradation of oxidant-damaged proteins. In contrast, immunoprecipitation of proteasomes with polyclonal or monoclonal antibodies prevented the ATP/ubiquitin-dependent degradation of lysozyme and also blocked the ATP-stimulated degradation of ubiquitin-conjugated lysozyme in reticulocyte and skeletal muscle extracts. These data indicate a critical role of the proteasome in the degradation of ubiquitin-conjugated proteins and suggest that the proteasome is associated with or is a component of the larger ubiquitin-conjugate-degrading enzyme complex.
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PMID:Involvement of the proteasome in various degradative processes in mammalian cells. 253 95

Tubulin can stimulate specifically the aryl phosphatase activity of the low-Mr polycation-stimulated (PCSL) phosphatase, measured as p-nitrophenyl phosphatase activity, or using reduced carboxamidomethylated and maleylated (RCM) lysozyme, phosphorylated on tyrosyl residues, as a substrate. This stimulation is independent of the degree of polymerization of tubulin (A50 = 60 nM) and is due to an increase in Vmax. It is mechanistically different from the ATP-induced activation and resistant to heat and trypsin treatment. Chymotrypsin destroys the stimulatory effect of tubulin. The polycation-stimulated phosphorylase phosphatase activity is inhibited by tubulin, probably by a polycation/polyanion interaction. The microtubule-associated protein, MAP2, is inhibitory to the p-nitrophenyl phosphatase activity and tubulin can eliminate this inhibitory effect. MAP2 also inhibits the polycation-stimulated phosphorylase phosphatase activity.
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PMID:Tubulin and MAP2 regulate the PCSL phosphatase activity. A possible new role for microtubular proteins. 254 1

Report of a case of multicentric reticulohistiocytosis with clinical, histopathologic, histochemical and immunohistochemical study. Neutral mucins and lipids were demonstrated in the cytoplasm of dermal histiocytes. The monocyte-histiocytic nature of the cells was shown by means of acid phosphatase, ATP-ase, muramidase and alpha-1-antitrypsin. Due to the immunohistochemical results, the relationship with Langerhans cells was rejected. No secretory activity of immunoglobulins was shown and the proliferation qualified as benign. The cells among the histiocytic population were typified as mature T lymphocytes.
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PMID:[Multicentric reticulohistiocytosis: histochemical, histoenzymatic and immunocytochemical study]. 269 57

Although protein breakdown in most cells seems to require metabolic energy, it has only been possible to establish a soluble ATP-dependent proteolytic system in extracts of reticulocytes and erythroleukemia cells. We have now succeeded in demonstrating in soluble extracts and more purified preparations from rabbit skeletal muscle a 12-fold stimulation by ATP of breakdown of endogenous proteins and a 6-fold stimulation of 125I-lysozyme degradation. However, it has still not been possible to demonstrate such large effects of ATP in similar preparations from liver. Nevertheless, after fractionation by DEAE-chromatography and gel filtration, we found that extracts from liver as well as muscle contain both the enzymes which conjugate ubiquitin to 125I-lysozyme and an enzyme which specifically degrades the ubiquitin-protein conjugates. When this proteolytic activity was recombined with the conjugating enzymes, ATP + ubiquitin-dependent degradation of many proteins was observed. This proteinase is unusually large, approx. 1500 kDa, requires ATP hydrolysis for activity and resembles the ubiquitin-protein-conjugate degrading activity isolated from reticulocytes. Thus the ATP + ubiquitin-dependent pathway is likely to be present in all mammalian cells, although certain tissues may contain inhibitory factors.
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PMID:Skeletal muscle and liver contain a soluble ATP + ubiquitin-dependent proteolytic system. 282 Mar 75


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