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

---

Query: EC:3.5.1.52 (PNGase F)
1,527 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A cholesteryl ester transfer protein (CETP) of apparent Mr 74,000 has recently been purified from human plasma. Cholesteryl ester transfer activity was found to accumulate in the medium of cultured Hep G2 cells. The transfer activity was removed by immunoprecipitation with specific antibodies to the plasma CETP. Sodium dodecyl sulfate gel electrophoresis of immunoprecipitates prepared from the medium of cells pulsed with [35S]methionine revealed a broad specific band of protein of Mr 72,000 to 76,000; by contrast, immunoprecipitates of cellular homogenates showed a sharp specific band of Mr 58,000. The Mr 72,000 to 76,000 band disappears, concomitant with the appearance of lower Mr products, upon neuraminidase or glycopeptidase F treatment of medium immunoprecipitates or of purified CETP. The results indicate that liver cells have the capacity to synthesize and secrete CETP. The CETP peptide acquires asparagine-linked carbohydrate and sialic acid during intracellular processing.
...
PMID:Cholesteryl ester transfer protein is secreted by Hep G2 cells and contains asparagine-linked carbohydrate and sialic acid. 331 17

Our laboratory recently reported the purification of a unique immunosuppressive glycoprotein isolated from human pregnancy urine (7). This glycoprotein, which we term uromodulin, has a m.w. of 85,000 as assessed on SDS-PAGE and is 30% carbohydrate. Uromodulin blocks in vitro antigen-specific T cell proliferation to recall antigens such as tetanus toxoid at concentrations as low as 100 pM. This glycoprotein also blocks the in vitro generation of spontaneous monocyte-mediated cytotoxicity (7, 36). Recent evidence strongly suggests that the primary action of uromodulin is to act as a specific ligand and modulator of IL 1 (10, 33). We now report additional biochemical characterization of uromodulin, and based on three independent lines of evidence, find that its immunologic activity appears to result from its glycosylation. First, measures to alter the tertiary folding of the protein backbone of uromodulin, including succinylation or reduction and carboxymethylation, fail to significantly affect its in vitro bioactivity. Second, after extensive digestion of intact uromodulin with pronase, the majority of the in vitro bioactivity can be recovered in a single carbohydrate-rich fraction. Finally, digestion with N-glycanase (N-glycosidase F-, an enzyme specific for N-asparagine-linked oligosaccharides) and subsequent purification on thin layer chromatography yields a single complex oligosaccharide that appears to be responsible for the majority of the in vitro immunosuppression mediated by uromodulin. These data suggest that uromodulin displays N-linked carbohydrate sequences capable of down-regulating antigen-specific T cell responses in vitro. It has been suggested that endogenous lectins may play an important role as recognition molecules in mammalian, as well as more primitive immune systems (23, 24). Our in vitro biologic data strongly suggest that the carbohydrate portion of uromodulin is an excellent candidate to function as a potential lectin receptor.
...
PMID:In vitro evidence that carbohydrate moieties derived from uromodulin, an 85,000 dalton immunosuppressive glycoprotein isolated from human pregnancy urine, are immunosuppressive in the absence of intact protein. 349 76

IgE-binding factors (IgE-BFs) were purified from the culture supernatant of RPMI-8866 cells, a human lymphoblastoid B-cell line expressing IgE receptors. The material, purified by affinity-chromatography on immunoadsorbents coupled to IgE or to monoclonal antibody against IgE receptor, was comprised of two major components with apparent molecular weight (MW) of 25,000-27,000 and 12,000, as determined by SDS-PAGE and silver staining. Only the 25,000-27,000 MW molecules were identified as IgE-BFs, as demonstrated by their reactivity with MabER in the Western blot and the immunoprecipitation assays, and their ability to inhibit rosette formation of U937 cells with IgE- but not with IgG-coated erythrocytes. IgE-BFs were purified to homogeneity by combining affinity-chromatography and either DEAE-ion exchange or reverse-phase chromatography on an HPLC system. Chromatofocusing analysis demonstrated the microheterogeneity of IgE-BFs that were comprised of molecules with isoelectric points ranging from 5.0 to 4.4. IgE-BFs were sensitive to treatment with O-glycosidase but not with N-glycanase. These molecules were resistant to heat and to pH ranging from 2 to 9; their immunoreactivity was lost after treatment with trypsin and pepsin. Papain digestion of purified IgE-BFs generated 14,000-16,000 MW molecules that were still binding to IgE and to MabER.
...
PMID:Purification and partial biochemical characterization of IgE-binding factors secreted by a human B lymphoblastoid cell line. 349 83

The rat major histocompatibility complex class I antigens RT1.Au and RT1.Eu from the u haplotype and RT1. An from the n haplotype were labeled with 14C-asparagine or with 3H-fucose, mannose, galactose, and N-acetylglucosamine. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis showed complete removal of radioactivity from the sugar-labeled antigen heavy chains by digestion with glycopeptidase F, an enzyme that removes N-linked glycans completely. High performance liquid chromatography analysis of the tryptic digests of the mixed sugar-labeled and asparagine-labeled antigens demonstrated that all the sugar-labeled peptides were coincident with asparagine-labeled peptides. The An antigen showed three glycopeptides, each of which had different amounts of sugar radioactivity. The antigens Au and Eu showed two glycopeptides with different amounts of radioactivity but at identical positions in the two antigens. Antigen Eu had an additional glycopeptide with a lower amount of radioactivity. The positions of the glycopeptides from the Au and Eu antigens were different from those of the An antigen. The peptide profiles of the 14C-asparagine-labeled Au and Eu antigens demonstrated distinct differences between the molecules. The results of this study show that: (a) all the glycans on rat class I antigens are N-linked, as they are on H-2 and HLA class I antigens; (b) there are compositional differences among the glycans in each of the three antigens; (c) the glycosylation pattern of the rat class I antigens is similar to that of the mouse class I antigens, which contain two or three glycans, in contrast to that of the human class I antigens, which contain only one glycan; and (d) the antigens Au and Eu from the same haplotype are more closely related to each other than they are to the An antigen.
...
PMID:Carbohydrate moieties of rat MHC class I antigens. 365 37

Lectins were isolated from fruits and leaves of Clerodendron trichotomum by affinity chromatography on lactamyl-Sepharose. The purified lectins (C. trichotomum agglutinin: CTA) were homogeneous on SDS/polyacrylamide gel electrophoresis, and the carbohydrate moiety was characterized by physicochemical and immunochemical methods. The asparagine-linked oligosaccharides were released by treatment with N-oligosaccharide glycopeptidase (almond, EC 3.5.1.52) of peptic glycopeptides obtained from fruit CTA, and separated by gel filtration and thin-layer chromatography. The structure of the predominant oligosaccharide was determined as Xyl beta 1----2 (Man alpha 1----6)(Man alpha 1----3)Man beta 1----4GlcNAc beta 1----4(Fuc alpha 1----3)GlcNAc by high-performance liquid chromatography, sugar analysis and 1H-NMR spectroscopy. The reactivity of the carbohydrate moiety of CTA toward various lectins was studied. Fruit and leaf CTAs were applied to polyacrylamide gel electrophoresis, transferred to nitrocellulose sheets and detected with horseradish-peroxidase-conjugated lectins. Concanavalin A, lentil lectin, pea lectin, Vicia faba lectin and Ulex europeus agglutinin I, but not wheat germ lectin, bound to fruit CTA. The results indicate new binding properties of these plant lectins: a beta-xylosyl residue substituted at C-2 of the beta-mannosyl residue of N-linked oligosaccharide does not affect the binding with mannose-specific lectins, lentil, pea and Vicia faba lectins can bind to N-linked oligosaccharides containing an alpha-L-fucosyl residue attached to C-3 of the asparagine-linked N-acetyl-D-glucosamine residue, and Ulex europeus agglutinin I can bind to the (alpha 1----3)-linked fucose residue of the N-linked oligosaccharide.
...
PMID:Characterization of the carbohydrate moiety of Clerodendron trichotomum lectins. Its structure and reactivity toward plant lectins. 379 15

A glycoprotein that circulates in human blood, binds to the surface of platelets and white cells and also binds serotonin with high affinity and specificity has previously been purified and partially characterized. This glycoprotein has been called serotonectin. Antibodies raised against serotonectin inhibited the uptake of [3H]serotonin by platelets. We now report on the amino acid and carbohydrate composition of this protein as well as on some of the properties of the protein from which the carbohydrate moiety was removed. Serotonectin (apparent molecular weight 200 000; as judged by SDS-polyacrylamide gel electrophoresis) is an acidic protein that contains about 13% carbohydrate (w/w) consisting of mannose, galactose, glucosamine and sialic acid in a molar ratio of 2:1:4:0.8. Initial characterization suggests that serotonectin is a sialoglycoprotein of complex-type oligosaccharide N-linked to asparagine through N-acetylglucosamine. Treatment of serotonectin with neuraminidase resulted in a quantitative release of sialic acid without loss of antigenicity or binding capacity for [3H]serotonin. Treatment of desialylated serotonectin under non-denaturing conditions with almond glycopeptidase A resulted in 60-80% release of sugar. The protein moiety of the glycopeptidase-digested material showed no change in the capacity to bind [3H]serotonin and exhibited the same antigenic properties as untreated serotonectin. These data show the non-involvement of the carbohydrate moiety of human serotonectin in the mechanism of binding serotonin but the possible contribution of this moiety to a tighter interaction with the serotonectin receptor.
...
PMID:Human serotonectin: a blood glycoprotein that binds serotonin. Chemical and physiological characterization. 394 85

Isolated plasma membranes from rat liver and ascites hepatoma cells were shown by SDS-polyacrylamide gel electrophoresis and concanavalin A reactivity to contain a variety of glycoproteins having asparagine-linked oligosaccharides. Membrane oligosaccharides were released by almond glycopeptidase digestion, and the pyridylamino derivatives were analyzed by high-performance liquid chromatography. Forty-four percent of the total carbohydrates in the original membranes were released and suggested to be of the complex type. Hepatoma membranes showed different oligosaccharide patterns from normal.
...
PMID:Analysis of asparagine-linked oligosaccharides from plasma membranes of rat normal liver and ascites hepatoma cells. 647 25

In order to obtain more information on the molecular structure of the angiotensin II (Ang II) binding sites from whole rat lung membranes these were characterized by isoelectric focusing (IEF) and SDS-PAGE. Whereas a single population of Ang II receptor sites was identified (Kd = 2.2 +/- 0.3 nmol/l; Bmax = 203.9 +/- 15.8 fmol/mg protein) by Scatchard analysis, using IEF three Ang II binding isoforms were observed; a major band which migrated to isoelectric point (pI) 6.7, and two minor bands with pI values of 6.5 and 6.3. Specific binding of 125I-Ang II to rat lung membrane preparations was sensitive to Losartan, a non-peptide AT1 receptor subtype antagonist, but was unaffected by the AT2 receptor subtype antagonist CGP42112A. Immunoblotting analyses on SDS gels, using a monoclonal antibody specific to the AT1 receptor, showed two immunoreactive protein species of 45 and 48 kDa. Enzymic deglycosylation using recombinant N-glycanase did not alter the molecular weight patterns of the AT1 receptor subtype. The results of the present study demonstrated that the Ang II receptor population in the whole rat lung consists solely of the AT1 receptor subtype and that the AT2 receptor subtype is absent. In addition, the data showed the existence of charge heterogeneity of the AT1 receptor subtype, and suggest that glycosylation probably does not contribute to its charge heterogeneity.
...
PMID:Charge heterogeneity of the AT1 angiotensin II receptor subtype in the rat lung. 749 May 29

In this study we demonstrate that the binding region of recombinant truncated human bone osteonectin (tHON) for type V collagen resides between amino acids 1 and 146. After removal of oligosaccharide chain structures from tHON, bovine bone osteonectin (BBON) and human platelet osteonectin (HPON) by N-glycanase, their ability to bind to type V collagen is increased, and HPON affinity to collagen V is the same as that of BBON. These data suggest that glycosylation of osteonectin has a direct or regulatory effect on osteonectin binding to collagen V and that the increase in tHON binding upon removal of carbohydrate is the result of a loss of a down-regulation site or direct interference of the carbohydrate at the binding site. To determine the specific role of each N-glycosylation site in tHON, Asn71 and Asn99 were mutated to Gln (N71Q, N99Q) and Thr73 and Thr101 mutated to Ala (T73A, T101A) to selectively inhibit oligosaccharide attachment. The binding affinity of N99Q and T101Q to collagen V is markedly increased over wild-type tHON, whereas N71Q and T73A are the same as wild-type tHON. The doubled mutant (N71,99Q) binds identically to collagen V as N99Q and T101A. These data suggest that only the position 99 glycosylation site (Asn99-X-Thr101) in tHON is important in the reduction of binding of osteonectin to collagen V. Consistent with the binding data is the observation that both the N71Q and T73A mutant proteins migrate on SDS-polyacrylamide gel electrophoresis gels identically to wild-type tHON, suggesting that there is little or no N-glycosylation of residue 71 in wild-type osteonectin.
...
PMID:Role of N-linked glycosylation in human osteonectin. Effect of carbohydrate removal by N-glycanase and site-directed mutagenesis on structure and binding of type V collagen. 755 69

Ammodytoxin A, the presynaptic neurotoxin from Vipera ammodytes ammodytes venom, was found to bind specifically and with high affinity to bovine cortex synaptic membrane preparation. The detected ammodytoxin A high-affinity binding was characterized by equilibrium binding analysis which revealed a single high-affinity binding site with Kd 4.13 nM and Bmax 6.67 pmoles/mg of membrane protein. 125I-ammodytoxin A was covalently cross-linked to its neuronal acceptor using a chemical cross-linking technique. As revealed by subsequent SDS-PAGE analysis and autoradiography, 125I-ammodytoxin A specifically attached to membrane components with apparent mol. wts 53,000-56,000. Besides by the native ammodytoxin A, the binding of radioiodinated ammodytoxin A to the neuronal acceptor was highly attenuated, also by other two iso-neurotoxins from V. a. ammodytes venom, ammodytoxins B and C, and neurotoxin crotoxin B from the venom of the South American rattlesnake (Crotalus durissus terrificus). Vipera berus berus phospholipase A2 was a weaker inhibitor, whereas nontoxic phospholipase A2, ammodytoxin I2 and myotoxic phospholipase A2 homologue, ammodytin L, both from V. a. ammodytes venom as well, were very weak inhibitors. No inhibitory effect on 125I-ammodytoxin A specific binding at all was, however, obtained with alpha-dendrotoxin, beta-bungarotoxin and crotoxin A, respectively. Treatment of synaptic membranes with proteinase K and Staphylococcus aureus V-8 proteinase, a combination of PNGase F and neuroaminidase, heat or acid lowered the 125I-ammodytoxin A specific binding to various extents but never completely abolished it. The ammodytoxin A binding site in bovine synaptic membranes is thus most likely a combination of membrane glycoprotein acceptor and membrane phospholipids. As ammodytoxin A reduced the second negative component of the perineural waveform, measured on mouse triangularis sterni preparation, which is very likely a result of an inhibition of a fraction of the terminal K+ currents, the ammodytoxin A acceptor could well be connected with K+ channels.
...
PMID:Ammodytoxin A acceptor in bovine brain synaptic membranes. 757 Jun 29


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>

---