Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.3.1 (alkaline phosphatase)
 47,916 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We compared sialidase (neuraminidase; EC 3.2.1.18) from Vibrio cholerae, Clostridium perfringens, and Arthrobacter ureafaciens, seeking to improve the electrophoretic separation of the liver and bone isoenzymes of alkaline phosphatase (EC 3.1.3.1) on cellulose acetate membranes. Resolution is decisively determined by the type and activity of sialidase used in the preincubation of serum sample. Sialidase from Arthrobacter ureafaciens is not suited for this method. For optimal separation of the two isoenzymes we recommend the use of sialidase from Vibrio cholerae, determination of its activity with a standard procedure such as described here (mucin or sialyl lactose as substrates), and a final concentration of sialidase activity of 2.0 or 2.9 U/L (measured with mucin or sialyl lactose) in the incubation mixture.
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PMID:Sialidase from different sources compared for electrophoretically separating serum alkaline phosphatase fractions from liver and bone. 277 24

Electrophoretic patterns of seminoma- and normal-testis-derived alkaline phosphatase isozymes, the placental alkaline phosphatase (PLAP)-like enzyme and the tissue-nonspecific (liver) alkaline phosphatase (LAP), were studied on starch gel and isoelectric focusing (IEF). Different migration patterns of the PLAP-like enzyme were observed with respect to both seminomas and normal testes on starch gel electrophoresis. On IEF, seminomas showed different staining patterns among different tumors; however, a common main activity was focused at pIs of 4.3-4.6, corresponding to pIs of PLAP. Normal testes showed two enzyme-staining regions, at pIs of 4.1 and 5.0-5.2, which were discriminated from pIs of PLAP and the PLAP-like enzyme in seminoma. The PLAP-like enzyme in seminoma was differentiated from PLAP by digestion with neuraminidase. Neuraminidase treatment simplified the distribution patterns of the PLAP-like enzyme in normal testis, but did not alter the pattern of microheterogeneity in seminoma. Two factors other than sialylation, namely structural modification of the carbohydrate moiety and variation of hydrophobicity, were shown to contribute to the microheterogeneity of the PLAP-like enzyme in seminoma. LAP in seminoma and in normal testis also showed marked electrophoretic heterogeneity and differences in pI distributions from LAP of liver. However, the migration patterns after desialylation were very similar to each other. The findings imply that electrophoretic heterogeneity demonstrated in LAP in seminoma and in normal testis is caused by a difference in sialic acid content in the molecule, and the heterogeneity of the PLAP-like enzyme in seminoma is considerable.
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PMID:Electrophoretic heterogeneity of alkaline phosphatase isozymes in seminoma and normal testis. 278 Dec 21

Isolation of two membrane-bound alkaline phosphatase (AP) species from avian growth plate cartilage matrix vesicle (MV) fractions is described. AP was first released from the membranes by phosphatidylinositol-specific phospholipase C (PIase C), followed by chromatography on DEAE-Bio-Gel A and Reactive-Red agarose. Two AP species having apparent Mr of 81.5 and 77 kDa by SDS-PAGE were purified in high yield and specific activity by this simple method. Treatment with neuraminidase to remove sialic acid residues reduced their size slightly, but did not diminish the difference in Mr between the two species. Digestion with N-glycanase, however, decreased both AP species to a common size of 59 kDa. This reveals that both enzymes are highly glycosylated and suggests that the two forms may result from differences in degree of glycation. The amino acid compositions of the two avian enzyme forms are very similar, but are markedly enriched in serine, glycine and glutamate when compared to those reported for mammalian liver-kidney-bone AP. Possible differences in amino acid sequence between the two avian forms have not been excluded. The cross-reactivity of polyclonal antibodies to these enzymes with bovine kidney, but not intestinal AP, indicate that the avian cartilage APs are of the liver-kidney-bone isozyme type.
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PMID:Isolation of two glycosylated forms of membrane-bound alkaline phosphatase from avian growth plate cartilage matrix vesicle-enriched microsomes. 280 49

Binding of two monoclonal anti-liposome antibodies to the surface of cultured murine peritoneal macrophages was investigated by indirect immunofluorescence and enzyme-linked immunosorbent assay. Neither antibody bound to cultures of freshly explanted, nonadherent macrophages, but immunoreactivity was observed following cell adherence to tissue culture plastic. Fluorescent microscopic evaluation revealed heterogeneity in staining patterns of the antibodies on adherent cells. Binding both to viable and fixed adherent macrophages was observed even after a 10,000-fold dilution of antibody. Treatment of adherent macrophage cultures with trypsin increased antibody binding. Further treatment of trypsinized-macrophages with alkaline phosphatase or neuraminidase did not affect antibody binding, but phospholipase D and, to a greater extent, phospholipase C resulted in a marked decrease in cellular binding. The data indicate that antibodies produced against liposomes appear to bind to surface phospholipids of macrophages, but binding can be influenced by the physiological state of the macrophage and overlying cell surface proteins.
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PMID:Antibodies to phospholipids and liposomes: binding of antibodies to cells. 282 Apr 89

Arylsulfatase A was purified from human lung and human placenta to apparent homogeneity presented by electrophoresis in the absence and presence of sodium dodecyl sulfate. The enzyme from normal lung, placenta, and lung adenocarcinoma showed considerable charge heterogeneity when examined by isoelectrofocusing, with isoelectric point (pI) ranging from 5.1 to 4.6. The enzyme from adenocarcinoma was more heterogeneous and having more acidic components than the other enzyme. When the tumor enzyme was treated with exogenous sialidase, alkaline phosphatase, or endo-beta-N-acetylhexosaminidase H (endoglycosidase H), the acidic components of the enzyme shifted to the more alkaline region on the focussing gel. The banding pattern of the enzyme from normal tissues also changed to the more alkaline region when treated with exogenous hydrolase and showed almost the same pattern as hydrolase treated enzyme from adenocarcinoma. Combined treatment of the enzyme with endoglycosidase H and sialidase resulted in complete loss of the most acidic components to give the less acidic components with pI of 5.1.50. and 4.9. Cyclic AMP-dependent protein kinase could not phosphorylate the protein moiety of arylsulfatase A even after the enzyme was treated with alkaline phosphatase. When an acidic fraction of the endoglycosidase H sensitive oligosaccharides from arylsulfatase A was treated with phosphatase, the acidic oligosaccharide fraction lost the negative charge on QAE-Sephadex chromatography. These results strongly suggest that the charge heterogeneity of arylsulfatase A is due not only to sialylation but also to phosphorylation at the carbohydrate moiety of the enzyme, and that the extent of substitution by acidic groups, sialic acid residue and phosphate residue, is markedly increased in the tumor enzyme.
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PMID:[Studies on charge heterogeneity of arylsulfatase A from human lung cancer]. 286 24

Treatment with cholecalciferol or 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) increases activity and changes electrophoretic mobility of alkaline phosphatase (alkPase) from duodenal brush border of vitamin D-deprived chicks. Three of the four molecular forms of the enzyme show reduced velocity of migration 9 h after 1,25(OH)2D3 or 24 h after vitamin D3. This change is reversed about 48 h later, when mobility of those bands is higher than that of controls. Incubation of enzyme preparations with exogenous neuraminidase produces the same electrophoretic modifications observed during the early stage, indicating that they are due to desialylation. Cholecalciferol or 1,25(OH)2D3 increase sialidase activity of duodenal brush border. This increment precedes that of alkPase and could account for the initial desialylation and moderate rise of alkPase. Cycloheximide markedly reduces alkPase in rachitic chicks and blocks the increase of the enzyme activity produced by vitamin D3, but does not modify the rise of sialidase or the reduction of alkPase electrophoretic mobility. The bimodal response of alkPase to 1,25(OH)2D3 or cholecalciferol comprises two different mechanisms: during a first stage, epigenetic modifications of preexisting enzyme can be triggered by the increased Ca2+ levels; in a second phase, there is activation of enzyme synthesis.
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PMID:Changes of intestinal alkaline phosphatase produced by cholecalciferol or 1,25-dihydroxyvitamin D3 in vitamin D-deficient chicks. 299 Mar 44

Gastric and intestinal phenotypic expression in 37 surgically obtained primary signet ring cell carcinomas, five of their metastases to lymph nodes, and three signet ring cell carcinomas transplanted into nude mice were determined by biochemical, mucin, histochemical, and ultrastructural studies. Crude extracts of cancer tissues were used for measurements of pepsinogen isozymes, sucrase, aminopeptidase (microsomal), and alkaline phosphatase. Histochemical staining of mucin by paradoxical concanavalin A, the galactose oxidase-Schiff sequence and sialidase-galactose oxidase-Schiff, and the periodate-borohydride technique/potassium hydroxide/periodic acid-Schiff procedure was performed. The procedures allowed clear definition of pyloric gland, surface mucous, small and large intestinal goblet, and intestinal absorptive cell types. Of 40 specimens examined, 19 consisted entirely of gastric-type cells, and three entirely of intestinal-type cells. The others consisted of mixtures of gastric and intestinal-type cells. The observed high incidence of intestinal-type cells in signet ring cell carcinomas suggested that intestinal-type cells develop independently from intestinal metaplasia within signet ring cell carcinomas (diffuse-type gastric cancers), which probably originate from nonmetaplastic gastric mucosa.
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PMID:Gastric and intestinal phenotypic expressions of human signet ring cell carcinomas revealed by their biochemistry, mucin histochemistry, and ultrastructure. 301

Two PRL-like glycoprotein hormone complexes were purified from the medium of cultured mouse conceptuses from day 10 of pregnancy: mouse placental lactogen-I (mPL-I) (29-32K), and mPL-I (36.5-42K). Sodium dodecyl sulfate-gel electrophoresis revealed that mPL-I (36.5-42K) is a complex of five proteins with mol wt of 36.5K, 37.5K, 39K, 40.5K, and 42K. Deglycosylation with peptide: N-glycosidase F or trifluoromethanesulfonic acid produced a single 29K protein. mPL-I (36.5-42K) was also sensitive to neuraminidase, but not to endo-beta-N-acetylglucosaminidase H or bacterial alkaline phosphatase. The production of intermediates from partial digestion of mPL-I (36.5-42K) with endo-beta-N-acetylglucosaminidase F indicated the presence of multiple glycosylation sites. mPL-I (29-32K) is a complex of three proteins with mol wt of 29K, 30.5K, and 32K. Treatment with peptide:N-glycosidase F or trifluoromethanesulfonic acid reduced the mol wt of the 30.5K and 32K bands to 28K. The 30.5K band was sensitive to endo-beta-N-acetylglucosaminidase H and endo-beta-N-acetylglucosaminidase F, but the 32K band was not. Neither band was sensitive to neuraminidase or bacterial alkaline phosphatase. The 29K band was resistant to all chemical and enzymatic treatments and is probably not glycosylated or phosphorylated. In the nonreduced state, neither form of mPL-I showed an increase in mobility over that of its reduced counterpart on sodium dodecyl sulfate-gel electrophoresis, indicating that neither form of mPL-I contains the large disulfide loop common to hormones of the PRL family. After iodination, all component proteins of both forms of mPL-I were found to bind to day 17 pregnant mouse liver membranes and were displaceable by excess mPL-II. In a radioreceptor assay, 125I-labeled mPL-I (36.5-42K) was displaced by mPRL or mPL-II, but not by mGH. An antiserum to both forms of mPL-I was generated, and a RIA employing mPL-I (36.5-42K) as the standard and radioligand was developed. Dilutions of day 10 pregnant maternal mouse serum and placental homogenate and a partially purified fraction of mPL-I (29-32K) produced displacement curves parallel to that of mPL-I (36.5-42K) standard curve. Five micrograms of mPRL, mPL-II, or mGH or 10 microliter day 17 pregnant or male mouse serum did not displace the radioligand from the antibody. mPL-I (36.5-42K) was lactogenic, but it did not possess LH-like bioactivity.
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PMID:Purification and partial characterization of two prolactin-like glycoprotein hormone complexes from the midpregnant mouse conceptus. 303 95

A modified agarose electrophoretic system for the separation of alkaline phosphatase (ALP, EC 3.1.3.1) isoenzymes is described. Bone, liver, high-molecular-mass, and intestinal ALP are separated with high reproducibility. The sensitivity of the agarose system is superior to cellulose acetate in detecting high-Mr ALP. Correlation is good between bone ALP fractions scanned before and after treatment with neuraminidase. Immunoglobulin-bound ALPs, the ALP-lipoprotein-X complex, and the additional ALP fraction observed in transient hyperphosphatasemia in children are detected by their peculiar electrophoretic mobility in the proposed system. Approximately 25% of the samples contained an additional fraction of intestinal-type ALP, as evidenced by neuraminidase treatment and use of polyclonal and monoclonal antibodies. Because the electrophoretic mobilities of this "intestinal variant" and of some immunoglobulin-bound ALP fractions are identical to those of bone and intestinal ALP, respectively, treatment of the samples with a polyclonal antibody that reacts with intestinal ALP is advised.
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PMID:Improved agarose electrophoretic method for separating alkaline phosphatase isoenzymes in serum. 304 80

A novel alkaline phosphatase (AP) isozyme was found in human adipose tissue. Adipose tissue alkaline phosphatase differed in enzymatic properties from liver, placental and intestinal alkaline phosphatases. On electrophoresis it showed the same mobility as intestinal alkaline phosphatase, but after treatment with neuraminidase its mobility was decreased to the same as or slightly less than that of neuraminidase-treated liver alkaline phosphatase. Its inhibition by amino acids, inactivation by urea and activation by Mg2+ were almost the same to those of liver alkaline phosphatase. However, at 56 and 65 degrees C it was more stable than liver alkaline phosphatase. Alkaline phosphatase activity was demonstrated histochemically in adipose tissue with naphthol AS-MX phosphate as substrate. It was localized in the wall of blood capillaries, but not present in adipocytes.
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PMID:A novel alkaline phosphatase isozyme in human adipose tissue. 310 Jan 9


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