Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:4.2.2.7 (heparinase)
 1,270 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Gene amplification of virus-specific sequences is widely used as a method to detect or confirm human immunodeficiency virus (HIV) infection. In this study we used an enzyme-linked affinity assay to quantify polymerase chain reaction products from whole blood, plasma, and separated mononuclear cells collected in the presence of four common anticoagulants: acid citrate dextrose, sodium EDTA, potassium oxalate, and sodium heparin. Attenuation of the product signal was observed after amplification of nucleic acid extraction from whole blood, washed mononuclear cells, and plasma from specimens collected in sodium heparin. These inhibitory effects on gene amplification could be reversed with heparinase. The addition of as little as 0.05 U of heparin completely inhibited amplification of an HLA-DQa sequence from placental DNA. We conclude that heparin can cause attenuation or inhibition of gene amplification. Acid citrate dextrose and EDTA, which lack inhibitory activity, are the most appropriate anticoagulants for clinical blood samples when polymerase chain reaction amplification is anticipated.
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PMID:Inhibition of human immunodeficiency virus gene amplification by heparin. 190 9

The human immunodeficiency virus-1 (HIV-1) Tat protein has previously been shown to transactivate the HIV-1-LTR when added exogenously to HeLa, H9 lymphocytic and U937 promonocytic cells growing in culture. Here we show that Tat enters these cells by adsorptive endocytosis. Tat appears to bind non-specifically to the cell surface, with greater than 10(7) sites per cell. A specific receptor was not detected by protein crosslinking experiments, and uptake was not affected by treating cells with trypsin, heparinase or neuraminidase. Uptake and transactivation could be inhibited by incubation with heparin, dextran sulfate, an anti-Tat monoclonal antibody, or by incubation at 4 degrees C. In contrast, transactivation by Tat was markedly stimulated by the addition of basic peptides, such as Tat 38-58 or protamine. Fluorescence experiments with rhodamine-conjugated Tat show punctate staining on the cell surface and then localization to the cytoplasm and nucleus. The lack of a specific receptor makes it unclear whether Tat uptake is biologically important in HIV infection, however, the efficiency of uptake raises the possibility that Tat may be useful for delivery of protein molecules into cells.
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PMID:Endocytosis and targeting of exogenous HIV-1 Tat protein. 205 Jan 10

The quantification of human immunodeficiency virus type 1 (HIV-1) RNA or hepatitis C virus (HCV) RNA has been facilitated by adapting a spin column procedure for sample preparation and the use of chemiluminescent detection of polymerase chain reaction (PCR) products in microtiter plate format. All materials were commercially available and relatively inexpensive. By making a single dilution prior to amplification, concentrations of 500 copies to 2.5 million HIV-1 1 RNA copies per mL and 1,000 copies to 50 million HCV RNA copies per mL could be determined on 140-microL samples. Between-run imprecision employing the improved procedure for HIV-1 RNA was 23%. Correlation of HIV-1 RNA concentrations obtained using chemiluminescent detection with values obtained by colorimetric assay of PCR products was 0.98. Correlation of HCV RNA concentration determined by the spin column-chemiluminescent assay procedure with those obtained by branched DNA methodology was 0.91. Spin columns could be used with serum or plasma containing acid-citrate-dextrose or heparin anticoagulant, but heparinized samples required treatment with heparinase prior to amplification.
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PMID:Improved methods for quantification of human immunodeficiency virus type 1 RNA and hepatitis C virus RNA in blood using spin column technology and chemiluminescent assays of PCR products. 898 50

Basic peptides such as human immunodeficiency virus type 1 (HIV-1) Tat-(48-60) and Drosophila Antennapedia-(43-58) have been reported to have a membrane permeability and a carrier function for intracellular protein delivery. We have shown that not only Tat-(48-60) but many arginine-rich peptides, including HIV-1 Rev-(34-50) and octaarginine (Arg(8)), efficiently translocated through the cell membranes and worked as protein carriers (Futaki, S., Suzuki, T., Ohashi, W., Yagami, T., Tanaka, S., Ueda, K., and Sugiura, Y. (2001) J. Biol. Chem. 276, 5836-5840). Quantification and time course analyses of the cellular uptake of the above peptides by mouse macrophage RAW264.7, human cervical carcinoma HeLa, and simian kidney COS-7 cells revealed that Rev-(34-50) and Arg(8) had a comparable translocation efficiency to Tat-(48-60). Internalization of Tat-(48-60) and Rev-(34-50) was saturable and inhibited by the excess addition of the other peptide. Typical endocytosis and metabolic inhibitors had little effect on the internalization. The uptake of these peptides was significantly inhibited in the presence of heparan sulfate or chondroitin sulfates A, B, and C. Treatment of the cells with the anti-heparan sulfate antibody or heparinase III also lowered the translocation of these peptides. These results strongly suggest that the arginine-rich basic peptides share a certain part of the internalization pathway.
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PMID:Possible existence of common internalization mechanisms among arginine-rich peptides. 1171 47

We hypothesize that in neurodegenerative disorders such as Alzheimer's disease and human immunodeficiency virus encephalitis the neuroprotective activity of fibroblast growth factor 1 (FGF1) against several neurotoxic agents might involve regulation of glycogen synthase kinase-3beta (GSK3beta), a pathway important in determining cell fate. In primary rat neuronal and HT22 cells, FGF1 promoted a time-dependent inactivation of GSK3beta by phosphorylation at serine 9. Blocking FGF1 receptors with heparinase reduced this effect. The effects of FGF1 on GSK3beta were dependent on phosphatidylinositol 3-kinase (PI3K)-protein kinase B (Akt) because inhibitors of this pathway or infection with dominant negative Akt adenovirus blocked inactivation. Furthermore, treatment of neuronal cells with FGF1 resulted in ERK-independent Akt phosphorylation and beta-catenin translocation into the nucleus. On the other hand, infection with wild-type GSK3beta recombinant adenovirus-associated virus increased activity of GSK3beta and cell death, both of which were reduced by FGF1 treatment. Moreover, FGF1 protection against glutamate toxicity was dependent on GSK3beta inactivation by the PI3K-Akt but was independent of ERK. Taken together these results suggest that neuroprotective effects of FGF1 might involve inactivation of GSK3beta by a pathway involving activation of the PI3K-Akt cascades.
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PMID:Fibroblast growth factor 1 regulates signaling via the glycogen synthase kinase-3beta pathway. Implications for neuroprotection. 1209 87

The hypervariable region 1 (HVR1) of hepatitis C virus (HCV) E2 envelope glycoprotein is a 27-amino-acid sequence located at its N terminus. In this study, we investigated the functional role of HVR1 for interaction with the mammalian cell surface. The C-terminal truncated E2 glycoprotein was appended to a transmembrane domain and cytoplasmic tail of vesicular stomatitis virus (VSV) G protein for generation of the chimeric E2-G gene construct. A deletion of the HVR1 sequence from E2 was created for the construction of E2DeltaHVR1-G. Pseudotype virus, generated separately by infection of a stable cell line expressing E2-G or E2DeltaHVR1-G with a temperature-sensitive mutant of VSV (VSVts045), displayed unique functional properties compared to VSVts045 as a negative control. Virus generated from E2DeltaHVR1-G had a reduced plaquing efficiency ( approximately 50%) in HepG2 cells compared to that for the E2-G virus. Cells prior treated with pronase (0.5 U/ml) displayed a complete inhibition of infectivity of the E2DeltaHVR1-G or E2-G pseudotypes, whereas heparinase I treatment (8 U/ml) of cells reduced 40% E2-G pseudotype virus titer only. E2DeltaHVR1-G pseudotypes were not sensitive to heparin (6 to 50 micro g/ml) as an inhibitor of plaque formation compared to the E2-G pseudotype virus. Although the HVR1 sequence itself does not match with the known heparin-binding domain, a synthetic peptide representing 27 amino acids of the E2 HVR1 displayed a strong affinity for heparin in an enzyme-linked immunosorbent assay. This binding was competitively inhibited by a peptide from the V3 loop of a human immunodeficiency virus glycoprotein subunit (gp120) known to bind with cell surface heparin. Taken together, our results suggest that the HVR1 of E2 glycoprotein binds to the cell surface proteoglycans and may facilitate virus-host interaction for replication cycle of HCV.
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PMID:The hypervariable region 1 of the E2 glycoprotein of hepatitis C virus binds to glycosaminoglycans, but this binding does not lead to infection in a pseudotype system. 1507 28

Dextran sulfate sodium (DSS) is a strong negatively charged heparin-like polysaccharide and has anti-immunodeficiency virus, anti-carcinogenesis, or occasionally tumor-promotion effects. The biological metabolism of DSS, however, remains unclear. In a previous study, we reported a novel method for the separation and quantification of DSS, using fluorometric labeling with 2-aminopyridine and a combination of size-exclusion and reverse phase high-performance liquid chromatography. In the present study, we have applied this method for analyses of in vitro chemical or enzymatic depolymerization of pyridylamino-DSS (PA-DSS). PA-DSS was depolymerized by specific enzymes such as alpha-amylase and alpha-glycosidase, but not by dextranase or heparinase. Unknown enzymes derived from cultured intestinal cells also strongly depolymerized PA-DSS as did alkaline substances. On the other hand, we have established a novel detection system using a post-column reaction. This method utilizes the spectrophotometrically metachromatic reaction of toluidine blue solution with DSS. This novel detection system may be specific and may potentially provide useful information in the analyses of sulfated polysaccharides, which are present in environmental and biological materials.
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PMID:The analysis of pyridylamino-dextran sulfate oligomers by high-performance liquid chromatography and a novel detection system for sulfated polysaccharides. 1525 3