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:3.1.3.1 (alkaline phosphatase)
47,916 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

An enzyme-linked lectin assay, termed ELLA, has been developed to investigate differences in cell surface carbohydrate groups on the surface of murine fibrosarcoma cells of varying metastatic potential. We have conjugated alkaline phosphatase to Griffonia simplicifolia I-B4 and have obtained an enzymatically-active probe for the detection of alpha-D-galactopyranosyl end groups. This probe has been used to detect different levels of expression of alpha-D-Galp end groups on the surface of unfixed murine tumor cell lines. Using the ELLA technique we have been able to demonstrate a correlation between the amount of these carbohydrate groups present on the cell surface and the degree of malignancy of the cell lines. Preliminary data suggest that the glycoprotein laminin accounts for a significant proportion of the cell surface alpha-D-Galp groups on these tumor cells and may play a functional role in their ability to metastasize. Furthermore, we have used ELLA to measure the binding of exogenous laminin to laminin-deficient cell lines and have found that this results in an increased ELLA reactivity. This increase corresponds to previously described increases in the malignant behavior of these cells after addition of exogenous laminin.
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PMID:A review of studies in our laboratory regarding ELLA methodology for the study of cell surface carbohydrates from tumors of varying metastatic potential. 390 Dec 26

Histopathologic and histochemical changes in experimental carcinomas following cryotreatment were observed to detect alkaline phosphatase (ALP) activity and UEA-1 lectin binding. Experimental carcinomas were induced in the hamster cheek pouch by topical application of 0.5% DMBA acetone solution twice a week. The cryoprobe at -60 degrees C was directly attached to the tumor surface for 90 sec. Histochemically, the tumor tissue following cryotreatment was completely destroyed in the surface area by the direct freezing and such cryonecrotic tumor tissue lacks stainability. Soon after cryotreatment and before cryonecrosis takes place, it has been observed that there is an intense dilatation of capillary vessels. Histochemically, high ALP activity was limited to capillary endothelium and to inflammatory cells. Lectin UEA-1 staining was usually found in both normal and neoplastic epithelial cells which were confined to capillary vessels. At 1 to 3 hr after cryotreatment, lectin UEA-1 binding was also positive in dilated endothelial cells of the frozen tissue as well as viable remaining neoplastic epithelia by freezing. ALP activity and UEA-1 binding disappeared in capillaries of cryonecrotic area in tumor tissues. Those findings suggest that biologic membrane changes in capillary endothelium of tumor stroma occur following cryotreatment.
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PMID:Identification of vascular system in experimental carcinoma for cryosurgery--histochemical observations of lectin UEA-1 and alkaline phosphatase activity in vascular endothelium. 392 63

The phosphorylation characteristics of insulin receptor from control and insulin-treated rat H-35 hepatoma cells 32P-labeled to equilibrium have been documented. The 32P-labeled insulin receptor is isolated by immunoprecipitation with patient-derived insulin receptor antibodies in the presence of phosphatase and protease inhibitors to preserve the native phosphorylation and structural characteristics of the receptor. The unstimulated insulin receptor contains predominantly [32P] phosphoserine and trace amounts of [32P]phosphothreonine in its beta subunit. In response to insulin, the insulin receptor beta subunit exhibits marked tyrosine phosphorylation and a 2-fold increase in total [32P]phosphoserine contents. High pressure liquid chromatography of the tryptic hydrolysates of the 32P-labeled receptor beta subunit from quiescent cells results in the resolution of up to 9 fractions containing [32P]phosphoserine. The insulin-stimulated tyrosine phosphorylation is concentrated in two of these receptor phosphopeptide fractions, whereas the increase in [32P]phosphoserine content is scattered in low abundance over all receptor tryptic fractions. Insulin receptors affinity-purified by lectin- and insulin-agarose chromatographies from insulin-treated, 32P-labeled cells exhibit a 22-fold increase in the Vmax of receptor tyrosine kinase activity toward histone when compared to controls. The elevated kinase activity of the insulin receptor derived from insulin-treated cells is not due to the presence of hormone bound to the receptor because the receptor kinase activity is assayed while immobilized on insulin-agarose. Furthermore, the insulin-activated receptor kinase activity is reversed following dephosphorylation of the receptor beta subunit with alkaline phosphatase in vitro. The correlation between the insulin-stimulated site specific tyrosine phosphorylation on receptor beta subunit and the elevation of receptor tyrosine kinase activity strongly suggests that the insulin receptor kinase is activated by hormone-stimulated autophosphorylation on tyrosine residues in intact cells, as previously demonstrated for the purified receptor.
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PMID:Tyrosine phosphorylation of insulin receptor beta subunit activates the receptor tyrosine kinase in intact H-35 hepatoma cells. 395 14

In a series of 46 cases of primary mammary ductal carcinoma, immunohistochemical markers of differentiation (casein, human placental lactogen, alphalactalbumin, pregnancy specific beta-1 glycoprotein, secretory component, CEA, and peanut lectin agglutinins [PLA]), were quantitated via point-counting. An immunoperoxidase bridge (PAP) was used to identify all except the PLA, in which an avidin-biotin complex with alkaline phosphatase development was employed. For none of the markers was there any difference in the quantity present in tumors of patients who had recurred versus the tumors of patients who had enjoyed a minimum of five years disease-free survival. Nonneoplastic epithelium was only rarely positive for these markers. Although eventually surmounted, technical problems significantly hampered application of morphometry to this histochemical material. The authors conclude that these markers have little relationship to differentiation toward mammary duct epithelium and that they do not provide significant prognostic information in patients with breast cancer.
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PMID:Correlation of immunohistochemical markers with patient prognosis in breast carcinoma: a quantitative study. 609 96

1. Proteins of fat-globule membrane from bovine milk were solubilized with the non-ionic detergent Triton X-100 in the presence of protease inhibitors. Approximately 25% of the total membrane protein was solubilized and the extracts were shown to contain a sample of most of the major membrane proteins and glycoproteins. 2. The solubilized proteins were separated in flat-beds of Ultrodex by electrofocusing and the pI values for the major proteins, glycoproteins and certain enzymes determined. Several of the proteins displayed marked heterogeneity indicating the existence of protein variants and isoenzymes. Principal pI values for the enzymes assayed were as follows: xanthine oxidase, 7.35--7.55; NADH2: iodonitrotetrazolium reductase, less than 4.5; 5'-nucleotidase, 7.15--7.4; alkaline phosphatase, 5.4--5.7; phosphodiesterase, 4.6--4.8; gamma-glutamyl transpeptidase, 4.4--4.55. 3. Fractions after electrofocusing were analyzed by 'fused rocket' immunoelectrophoresis and crossed immunoelectrophoresis after separation in polyacrylamide gels containing sodium dodecyl sulphate. Major antigens of the membrane include xanthine oxidase and glycoproteins of apparent molecular weights 67 000, 49 500 and 46 000. The latter two components share common antigenic determinants and could not be separated by gel filtration, ion-exchange chromatography, lectin-affinity chromatography or preparative electrofocusing.
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PMID:Separation of the proteins of bovine milk-fat-globule membrane by electrofocusing with retention of enzymatic and immunological activity. 610 13

We estimated the concentrations, multiple forms, and lectin binding of five microsomal enzymes in particle free extracts from human kidney, pancreas, jejunal mucosa, and normal and cancerous liver. While arylesterase markedly reacted only with concanavalin A, arylamidase, alkaline phosphatase, gamma-glutamyltransferase, and cholinesterase were intensely precipitated by lectins from Ricinus communis 120, Canavalia ensiformis, Triticum vulgare and Phaseolus vulgaris S. Agglutinins from Glycine max, Arachis hypogaea and Ulex europaeus proved less effective. The reaction mainly depended on the origin of enzymes not on their species. Desialylation always decreased precipitation, and in extracts of normal liver parenchyma it even totally abolished precipitation, by Triticum vulgare lectin. Sialoenzymes therefore appear to be normal intracellular constituents. Differences between enzymes from normal and cancerous liver were not reflected by variant properties of the corresponding activities in sera. The same held true for multiple forms. The reasons for these differences are discussed.
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PMID:[Catalytic concentration, multiple forms, and lectin affinity of microsomal enzymes from human tissues: lectins as reagents, II (author's transl)]. 612 Feb 6

Histochemical techniques have been applied to the identification of cell types cultured from human endometrium. Previous work from this laboratory characterized two principal cell types found in cultures of endometrium: a mature epithelial cell and another cell which was classified as the endometrial stromal cell based on light and electron microscopy. In this report we compare the histochemical staining of endometrial tissue in frozen sections to that of cultured cells. These results confirm the epithelial and stromal nature of the respective cell types. Several markers were found that could distinguish between cells of epithelial and stromal origin. The enzymes alkaline phosphatase, gamma-glutamyltranspeptidase, peroxidase, and beta-glucuronidase were localized in glandular and surface epithelia in frozen sections and in colonies of epithelial cells in culture. Stroma in frozen sections and cultured stromal cells contained leucine aminopeptidase and fibronectin. Epithelia in sections and in culture could also be distinguished from cells of stromal origin by preferential binding of lotus and peanut lectin. Several other markers were found in both endometrial epithelium and stroma.
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PMID:Histochemical identification of cultured cells from human endometrium. 614 95

Purpura was grossly observable in albino mice 6 to 8 h after the intraperitoneal injection of sterile, deoxyribonuclease-treated, cell-free extracts prepared by sodium deoxycholate-induced lysis, sonic disruption, Parr bomb treatment, autolysis without sodium deoxycholate, or alternate freezing and thawing of washed suspensions of Streptococcus pneumoniae type I. Cell-free extracts obtained from sonically disrupted, heat-killed cells (100 degrees C for 20 min) did not contain purpurogenic activity. The reaction was maximal at approximately 24 h postinjection, started to fade slowly after 24 to 48 h, and usually was not grossly observable by 4 to 6 days postinjection. The purpura-producing principle (PPP) in the cell-free extract was purified by sequential ammonium sulfate precipitation, protamine sulfate precipitation, Sepharose 6B gel filtration, wheat germ lectin-Sepharose 6MB affinity chromatography, ribonuclease and trypsin treatment, and a second Sepharose 6B gel filtration step. The final preparation (i) contained glucosamine (5.6%), muramic acid (8.0%), neutral carbohydrate (12.8%), phosphate (8.0%), orcinol-reactive material (6.0%), and Lowry-reactive material (1.6%), and (ii) was free of detectable amounts of deoxyribonucleic acid, capsular polysaccharide, neuraminidase, cytolysin, and hyaluronidase. The isoelectric point and molecular size of the PPP were approximately pI 3.0 and several million daltons, respectively, and the activity remained in the supernatant fluid after centrifugation for 1 day at 105,000 x g. PPP activity was destroyed by incubation with egg white lysozyme and sodium metaperiodate but was resistant to trypsin, pronase, alpha-amylase, deoxyribonuclease, ribonuclease, alkaline phosphatase, pancreatic lipase, 7% trichloroacetic acid, 6 M urea, autoclaving (121 degrees C) for 30 min, and mild acid and alkali exposure. Our observations indicate that the PPP requires intact beta-1,4-glucosidic linkages for activity and support the working hypothesis that activity is associated with pneumococcal peptidoglycan solubilized by the bacterium's autolysin.
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PMID:Characterization of pneumococcal purpura-producing principle. 624 53

Plasma membrane extracts from Herpes simplex virus type 1 transformed hamster embryo fibroblasts were chromatographed on Lens culinaris lectin coupled to Sepharose (LcH-Sepharose) and analysed by dodecyl sulphate polyacrylamide gel electrophoresis. Coomassie blue-staining revealed two major protein bands with apparent molecular weights of 125 000 and of about 75 000-90 000. In plasma membranes isolated from these tumor cells prior labeled with [3H]fucose or [3H]glucosamine these bands contained the highest amounts of incorporated radioactivity. Separation by LcH-Sepharose-affinity chromatography as well as metabolic labeling clearly demonstrates their glycoprotein character. The 125 000 protein coincides with alkaline phosphodiesterase I activity with a Km of 6 . 10(-4) M for TMP p-nitrophenyl ester and is competitively inhibited by UDP-N-acetylglucosamine. This enzymatic activity is also present in normal hamster embryo fibroblasts. Gel electrophoresis of the Lens culinaris lectin-binding glycoproteins from plasma membranes of normal hamster embryo fibroblasts additionally revealed a strong alkaline phosphatase activity represented by an apparent molecular weight of 150 000, while HSV1 hamster tumor cells contain only a very weak activity of this enzyme activity. HSV-lytically infected cells, however, have unchanged levels of alkaline phosphatase activity, whereas alkaline phosphodiesterase activity increases slightly.
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PMID:Alkaline phosphodiesterase I and alkaline phosphatase I in plasma membranes of herpes simplex virus type 1 transformed hamster cells. 627 77

In solubilized, (wheat germ) lectin-purified preparations of rat liver membranes, insulin stimulated the incorporation of 32P from [gamma-32P]ATP into tyrosine residues of insulin receptor, casein, and histones. Despite the presence of both protein kinase and phosphatase activities in these preparations, no decrease in the 32P content of receptors (preincubated with or without insulin (0.5-100 nM)) was detected whether 32P incorporation was terminated by excess ATP, ATP + Mn2+, EDTA, or phosphatase inhibitors. Similarly, there was no decrease in the 32P content of phosphoreceptors incubated for up to 60 min with fresh receptor preparations in the presence or absence of insulin. Dephosphorylation of the insulin receptor to 20% of original 32P content only occurred when alkaline phosphatase was added to the preparations. It is concluded that endogenous receptor phosphatase(s) are either missing or inactive in these preparations, and consequently, insulin stimulates phosphorylation of its own receptor by activating a protein kinase. The kinase activity is tightly associated with the receptor itself; insulin also stimulated the phosphorylation of both receptor subunits in purified insulin-receptor complexes that had been immunoprecipitated by anti-insulin antibodies. However, the phosphorylating machinery is much more sensitive to heat inactivation than the binding function (90% less 32P incorporation versus 15% less binding during 60-min incubation at 37 degrees C), suggesting that the kinase is not associated exclusively with the insulin-binding domain.
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PMID:Insulin stimulated phosphorylation of its own receptor. Activation of a tyrosine-specific protein kinase that is tightly associated with the receptor. 633 86


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