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)

The detection of human immunodeficiency virus (HIV)-associated antigens was simplified by the application of dot immunobinding on a nitrocellulose matrix. Antigens were detected by applying the polyethylene glycol-precipitated supernatants of experimentally infected cultures directly onto nitrocellulose strips and sequentially incubating the strips with an anti-HIV antiserum and an alkaline phosphatase-conjugated, species-specific antiserum. The immune reaction was developed by adding the precipitable substrate indoyl phosphate. The dot immunobinding assay was nearly as sensitive as the reverse transcriptase assay in detecting HIV antigens in experimentally infected peripheral blood mononuclear cells, as well as in a T-cell line. The technique was also useful in the in vitro evaluation of antiviral agents. The dot immunobinding assay is a simple and sensitive technique that is useful in the detection of HIV antigens in studies of viral pathogenesis.
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PMID:Dot immunobinding assay for detection of human immunodeficiency virus-associated antigens. 331 91

NMRI mice immunosuppressed with dexamethasone followed by challenge intraesophageally with axenic Giardia lamblia (Portland I) trophozoites had severe infection in terms of the trophozoite counts in the jejunum. Although the immunosuppressive treatment with cortisone itself resulted in a deleterious effect on brush border membrane enzymes, the decline in disaccharidases (sucrase, maltase, and lactase) and alkaline phosphatase was highly significant (P less than 0.001) following G. lamblia infection. The alterations in enzymatic activity in immune intact but infected animals demonstrated the potential of the parasite itself to cause damage to the brush border membrane. We believe that individuals with underlying immunodeficiency, upon infection with G. lamblia, may have increased damage of the brush border membrane, leading to severe malabsorption.
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PMID:Giardia lamblia infection in immunosuppressed animals causes severe alterations to brush border membrane enzymes. 276 19

Simple and sensitive methods to directly detect the human immunodeficiency virus (HIV) are needed for routine use in the clinical laboratory. In this study, we compared DNA probes prepared by: (1) nick translation with biotinylated dATP; (2) direct covalent biotinylation with photobiotin; (3) direct covalent reaction with 2-acetylaminofluorene (AAF); and (4) a standard radioactive (32P) nick translation procedure. These four DNA probes were hybridized with dilutions of purified target HIV DNA blotted onto nitrocellulose strips. Hybridization was detected using a complex of strepavidin-alkaline phosphatase [for (1) and (2)], alkaline phosphatase-tagged antibodies [for (3)] and by autoradiography [for (4)]. Alkaline phosphatase was detected colorimetrically using nitroblue tetrazolium and 5-bromo-4-chloro-3-indolyl phosphate. After 1 h, AAF probes were most sensitive (amount detected less than 5 pg), followed by biotin (10 pg), photobiotinylated probes (20 pg) and the radioactive probe (10 pg). The AAF probes were then used to detect HIV DNA in infected CEM cells. We conclude that non-radioactive DNA labelling methods can be used to directly detect HIV DNA under conditions compatible with present clinical laboratory procedures.
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PMID:Comparison of non-radioactive DNA hybridization probes to detect human immunodeficiency virus nucleic acid. 345 25

Jejunal biopsies from six patients having the small bowel enteropathy associated with common variable immunodeficiency have been subjected to analytical subcellular fractionation and enzymic and regulatory peptide microassay to define the organelle pathology of this syndrome. Compared with normal subjects, the immunodeficient patients had decreased activities of the three brush border enzymes: alkaline phosphatase, gamma-glutamyl transferase and alpha-glucosidase. The other organelle marker enzyme activities and all the regulatory peptide concentrations did not differ from the controls. Density gradient experiments showed a complete loss of particulate beta-glucosidase (lactase) with activity entirely located in the cytosol. The integrity of other organelles was normal. These data indicate that the enteropathy of common variable immunodeficiency is associated with abnormalities in the jejunal brush border analogous to those present in tropical malabsorption syndrome.
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PMID:Jejunal mucosal enzyme activities, regulatory peptides and organelle pathology of the enteropathy of common variable immunodeficiency. 369 46

Morphological studies of the thymus and the spleen were made in 54 alcohol addicts and 53 cases of other death cause at the age from 25 to 65 yr. Alcoholism was found to induce the reduction in thymus size and the number of its lobes with a simultaneous decrease of lymphocyte content in them; in the lymphatic follicles of the spleen (in the presence of infectious processes) absence of bright multiplication centres and alkaline phosphatase activity in their mantle zone cells was noted. There appeared to be a statistically significant difference in the amount of the parenchyma in the thymus of alcoholics (p less than 0.017) compared to that of the control group 0.68 +/- 0.09 g and 1.33 +/- 0.21 g, respectively and this figure was almost 4-fold less, than the average age norms, 0.26 +/- 0.03 g; the quantitative content of the splenic pulp in alcoholics also had a statistically significant difference compared to the control group (P less than 0.001), 6.37 +/- 0.66 g vs. 12.59 +/- 0.98 g, respectively. The data obtained indicate the development of immunodeficiency states in alcoholism and basing on the changes occurring in the thymus and spleen they can be referred to a combined T- and B-cell form of immunodeficiency.
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PMID:[Morphologic characteristics of the changes in the thymus and spleen in alcoholism]. 380 Jun 80

The activity of plasma membrane marker enzymes which are involved in purine metabolism (5'-nucleotidase, alkaline 5'-nucleotide phosphodiesterase), in active ion transport (Na-K-Mg-adenosine triphosphatase, ouabain-sensitive Na-K-adenosine triphosphatase), in aminoacid transport (gamma-glutamyltranspeptidase), and in basic physiologic functions (alkaline phosphomonoesterase) were assayed in mononuclear cells isolated from peripheral blood of normal donors and of patients with primary immunodeficiency. Irrespective of the clinical classification of the immunodeficiency, the cells of patients were characterized by significantly diminished 5'-nucleotidase and to a certain extent by lower alkaline phosphomonoesterase activities. Average activity levels of other enzymes were similar in cells of patients and controls, but scattering was more pronounced in the first group. Determination of substrate affinity revealed different kinetic properties of 5'-nucleotidase in cells from patients and normal donors; however, the extent of inhibition by beta-glycerophosphate or alpha, beta-adenosine-methylene diphosphate was comparable for both types of cells. The presence of inhibitory compounds in patients' serum was excluded by mixing experiments. When activities of the various plasma-membrane-associated enzymes were compared with each other, significant correlations emerged in normal lymphocytes. Most of these correlations were absent in cell membranes of immunodeficient patients. The findings indicate that the plasma membrane of lymphocytes from patients with immunodeficiency may be characterized by an altered distribution of enzymatic constituents.
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PMID:Correlations between enzymatic and immunologic properties of human peripheral blood mononuclear cells. I. Ectoenzymes of normal and immunodeficient peripheral blood mononuclear cells. 612 61

Plasmids were constructed whereby the expression of a reporter gene, either the cDNA corresponding to the secreted form of human alkaline phosphatase (SEAP) or the herpes simplex virus type 1 (HSV1) thymidine kinase (tk) gene, was rendered dependent upon the expression of the human immunodeficiency virus type 1 (HIV1) tat and rev proteins. The SEAP or tk genes were placed between HIV1 splice donor and acceptor sites. One SEAP construct carried a series of alternating splice donor and acceptor sites. In all cases, the rev response element mapped within an intron. Despite such mimicry of the HIV1 genome, residual expression of the reporter gene in the absence of tat and rev was observed. These results, as well as non-specific T-cell recruitment, suggest limits to the specificity of using HIV-activated toxic gene expression to kill HIV-infected cells.
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PMID:Residual expression of reporter genes in constructs mimicking HIV genome organization. 748 Oct 89

Monoclonal antibodies (MAbs) were raised against the gag protein of human immunodeficiency virus type 1 (HIV-1). The reactivity of the selected Mabs with the matrix protein p17 of HIV-1 were investigated in several tests, i.e. ELISA, immunostaining of Western blots, and alkaline phosphatase anti-alkaline phosphatase immunocytochemistry (APAAP). All three Mabs reacted exclusively with HIV-1 and showed specific binding to the virus surface in pre-embedding immuno-electron-microscopy and useful as diagnostic agents. In an "in vitro" cultivation experiment the MAbs showed antiviral activity in concentrations in the range of 25-100 micrograms/ml. No binding region could be defined using overlapping peptides consisting of the p17 protein sequence of HIV-1 in an epitope mapping system and therefore we concluded, that the MAbs recognize a conformational epitope.
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PMID:Characterization of murine monoclonal antibodies directed against the submembrane protein p17 of HIV-1. 750 41

A new method is described which is suitable for reliably analysing apoptotic fragmentation in small amounts of DNA. After isolation, DNA was labelled with biotin-4-dUTP using Klenow polymerase. Then DNA was size-separated by agarose gel electrophoresis, blot transferred and subsequently visualized by the streptavidin alkaline phosphatase-BCIP/NBT procedure. This non-radioactive method was used to detect apoptotic DNA in rat pheochromocytoma PC12 cells, treated with tributyltin (1 nM). While only 30 ng of DNA is required for analysis of apoptotic DNA using the new blot technique, 100-fold more material is needed to identify the fragmentation of DNA after separation by agarose gel electrophoresis and direct staining with ethidium bromide. In a further set of experiments, rat cortical cells were incubated with human immunodeficiency virus type 1 viral glycoprotein of M(r) of 120 kDa (gp120) to induce apoptosis. More than 0.3 ng of gp 120/ml are required to detect apoptotic DNA by the direct procedure; only 0.1 ng gp120/ml or less were sufficient to document clear DNA fragmentation using the non-radioactive blotting technique described here. These results demonstrate that the new procedure can be used to analyse very small amounts of apoptotic DNA and shows that gp120-induced apoptosis can be measured at low concentrations of the viral protein.
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PMID:A non-radioactive, sensitive method for the detection of DNA fragmentation in apoptotic cells (rat pheochromocytoma PC12 and rat cortical cells). 752 53

Binding and signaling proteins based on Escherichia coli alkaline phosphatase (AP; EC 3.1.3.1) were designed for the detection of antibodies. Hybrid proteins were constructed by using wild-type AP and point mutants of AP [Asp-101 --> Ser (D101S) and Asp-153 --> Gly (D153G)]. The binding function of the hybrid proteins is provided by a peptide epitope inserted between amino acids 407 and 408 in AP. Binding of anti-epitope antibodies to the hybrid proteins modulates the enzyme activity of the hybrids; upon antibody binding, enzyme activity can increase to as much as 300% of the level of activity in the absence of antibody or can decrease as much as 40%, depending on the presence or absence of the point mutations in AP. The fact that modulation is altered from inhibition to activation by single amino acid changes in the active site of AP suggests that the mechanism for modulation is due to structural alterations upon antibody binding. Modulation is a general phenomenon. The properties of the system are demonstrated by using two epitopes, one from the V3 loop of human immunodeficiency virus type 1 gp120 protein and one from hepatitis C virus core protein, and corresponding monoclonal antibodies. The trend of modulation is consistent for all hybrids; those in wild-type AP are inhibited by antibody, while those in the AP mutants are activated by antibody. This demonstrates that modulation of enzyme activity of the AP-epitope hybrid proteins is not specific to either a particular epitope sequence or a particular antibody-epitope combination.
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PMID:A molecular sensor system based on genetically engineered alkaline phosphatase. 754 Nov 35


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