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:3.2.1.23 (beta-galactosidase)
14,648 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The present study deals with the biochemical properties of high-molecular-weight glycopeptides isolated from the surface of human teratocarcinoma cells. This cell surface material released by mild trypsin digestion from galactose-labeled human teratocarcinoma cells, Tera I and PA1, was digested extensively with pronase. Most of the resulting glycopeptides were large and were excluded from a Sephadex G-50 column. The properties of these large cell surface glycopeptides isolated from Tera I cells have been examined in detail. It is clear from these experiments that they are neither acidic mucopolysaccharides nor mucin-type glycopeptides with short oligosaccharide chains. Although the glycopeptides are hardly hydrolyzed by beta-galactosidase even after prior digestion with neuraminidase, around 30% of the glycopeptides are depolymerized by treatment with endo-beta-galactosidase from Escherichia freundii. The large cell surface glycopeptides from Tera I cells therefore appear to be very similar to the large glycopeptides seen on mouse embryonal carcinoma cells, which have core structures composed of galactose and N-acetylglucosamine. Like the mouse cell glycopeptides, a fraction of the large glycopeptides from these human cells bind to agarose-conjugated fucose-binding proteins and peanut agglutinin.
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PMID:Biochemical properties of the high-molecular-weight glycopeptides released from the cell surface of human teratocarcinoma cells. 680 82

We have characterized the UDP-galactose: alpha-N-acetylgalactosaminide beta 3 galactosyltransferase in human tracheal epithelium using asialo ovine submaxillary mucin as the acceptor. Maximal enzyme activity was obtained at pH 6.0-7.5 and at 20-25 mM MnCl2 and at 2% Triton X-100. Cd2+ could substitute for Mn2+ as the divalent ion cofactor. Spermine, spermidine, putrecine, cadaverine, and poly-L-lysine stimulated the enzyme activity at low (2.5 mM) MnCl2 concentration. The apparent Michaelis constants for N-acetylgalactosamine, asialo ovine submaxillary mucin, and UDP-galactose were 15.5, 1.14, and 1.36 mM, respectively. The enzyme activity was not affected by alpha-lactalbumin. The alpha-N-acetygalactosaminide beta 3 galactosyltransferase was shown to be different from the N-acetylglucosamine galactosyltransferase by acceptor competition studies. The product of galactosyltransferase was identified as Gal beta 1 leads to 3GalNAc alpha Ser (Thr) by (a) isolation of [14C]Gal-GalNAc-H2 after alkaline borohydride treatment of the 14C-labeled product, (b) establishment of the beta-configuration of the newly synthesized glycosidic bond by its complete cleavage by bovine testicular beta-galactosidase, and (c) assignment of the 1 leads to 3 linkage by identification of threosaminitol obtained from the oxidation of the disaccharide with periodic acid followed by reduction with sodium borohydride, hydrolysis in 4 N HCl, and analysis on an amino acid analyzer. The 1 leads to 3 linkage was confirmed by its resistance to jack bean beta-galactosidase and by the presence of a m/e 307 ion fragment and the absence of a m/e 276 ion by gas-liquid chromatography-mass spectrometry analysis. When acid and beta-galactosidase-treated human tracheobronchial mucin was used as the acceptor, 3.3% of the product was found as [14C]Gal-GalNAc-H2. The remainder of the [14C]Gal was found in longer oligosaccharides formed by a different beta-galactosyltransferase. This galactosyltransferase is slightly inhibited by alpha-lactalbumin and stimulated by spermine.
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PMID:Mucin biosynthesis. Characterization of UDP-galactose: alpha-N-acetylgalactosaminide beta 3 galactosyltransferase from human tracheal epithelium. 680 62

A recently identified, high m.w. human tumor antigen, reactive with monoclonal antibody (MAb) PD41 and designated prostate mucin antigen (PMA), was found to have expression highly restricted to prostate carcinomas. Both biochemical and immunological characteristics of this antigen and its relationship to other tumor-associated mucins and various species of submaxillary mucins were evaluated. Immunohistochemical examination of submaxillary tissues revealed differential expression of this antigen and other human tumor-associated mucins, with MAb PD41 expression found only in bovine submaxillary gland serous cells. Neuraminidase treatment enhanced antibody binding by 50%, suggesting partial masking of the PD41 epitope by sialic acid. Antigenicity was reduced by treatment with alkaline-borohydride, sodium metaperiodate, beta-galactosidase, O-glycanase, and various proteolytic enzymes. MAb PD41 antibody binding also could be significantly reduced by selected lectins and sugars suggesting that the immunodominant carbohydrate involved in the epitope was an O-linked oligosaccharide containing N-acetyl galactosamine as a major component. This antigen was further distinguished from T, Tn, or Sialyl-Tn antigens and blood group carbohydrate antigens by radioimmunometric analyses. Cross-blocking and double determinant RIA experiments also showed a distinction between the PD41 epitope and several well-characterized tumor-associated mucin antigens such as MUCI, CEA, M344, OCI25, and AR3 as well as bovine submaxillary core protein. Our results indicate that the PD41-reactive epitope is a non-acidic O-linked carbohydrate or glycopeptide epitope with restricted expression in prostate carcinomas and bovine submaxillary glands. This expression is distinct from other mucin-like tumor-associated antigens identified in human prostate carcinomas.
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PMID:Characterization of a prostate carcinoma mucin-like antigen (PMA). 755 18

Glycophorin A was digested with glycoprotease (Pasteurella haemolytica) and the digest was fractionated by a combination of high-pressure column chromatographies to produce the glycopeptides GPA-1 to GPA-6. Sequence analysis of the glycopeptides revealed that two serine residues (Ser-14 and Ser-15) are not glycosylated, Thr-17 and Ser-19 being glycosylated instead, in disagreement with the accepted structure. The glycopeptides thus obtained were treated with sialidase and beta-galactosidase. The Tn antigenicity, as assayed by the binding to a monoclonal anti-Tn antibody (MLS 128), was found exclusively in the glycopeptides including three (cluster I) or four (cluster II) consecutive residues of GalNAc-Ser/Thr, whereas the glycopeptide (GPA-2) containing two nonconsecutive GalNAc-Ser/Thr residues had practically no Tn antigenicity. The immunoreactivities of GPA-1 and GPA-3, containing both clusters I and II, and GPA-4, containing cluster II, were 63% (calcd. 67%), 81% (calcd. 86%), and 50% (calcd. 50%), respectively, of the immunoreactivity of GPA-5 or GPA-6, containing cluster I (the average being taken as the basis), based on the reactivity per GalNAc residue. These results indicate that clusters I and II react with the antibody to the same extent. The structure consisting of three consecutive glycosylated Ser/Thr residues may be essential for Tn antigenicity in the light of previous results for ovine submaxillary mucin.
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PMID:Epitopic structure of Tn glycophorin A for an anti-Tn antibody (MLS 128). 768 97

The mucin-type carbohydrate Tn cryptantigen (GalNAc alpha 1-O-Ser/Thr, where GalNAc is N-acetyl-D-galactosamine) is expressed in many carcinomas, in haemopoietic disorders including the Tn syndrome, and on human immunodeficiency virus (HIV) coat glycoproteins, but is not expressed on normal, differentiated cells because of the expression of a Tn-processing galactosyltransferase. Using Jurkat T leukaemic cells which express high levels of Tn antigen due to deficient Tn galactosylation, we have established the Tn antigen-mediated gene transfer and demonstrate the considerable efficiency of this approach. We used poly(L-lysine) conjugates of the monoclonal antibody 1E3 directed against the Tn antigen to deliver the luciferase and beta-galactosidase reporter genes to Jurkat cells by receptor-mediated endocytosis. Addition of unconjugated 1E3 reduced transfection efficiency in a concentration-dependent manner and incubation with free GalNAc abolished DNA transfer completely, indicating that gene delivery is indeed mediated by the Tn antigen. Pre-treatment of Jurkat cells with Vibrio cholerae sialidase, which uncovers additional Tn antigens, resulted in an improvement of gene transfection. Both human and chicken adenovirus particles attached to the DNA/polylysine complex strongly augmented transgene expression. When the beta-galactosidase (lacZ) gene was delivered to Jurkat cells by Tn-mediated endocytosis, up to 60% of the cells were positive in the cytochemical stain using 5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside (X-gal) as a chromogenic substrate. The efficiency of the transferrin receptor-mediated DNA uptake into Jurkat cells was comparatively low, although these cells were shown to express considerable amounts of transferrin receptor. We show here that a mucin-type carbohydrate antigen mediates highly efficient DNA uptake by endocytosis into Jurkat T cells. This method represents a 50-fold improvement of Jurkat cell transfection efficiency over other physical gene transfer techniques. Specific gene delivery to primary cancer cells exhibiting Tn epitopes may especially be desirable in immunotherapy protocols.
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PMID:Carbohydrate receptor-mediated gene transfer to human T leukaemic cells. 782 4

A cDNA clone encoding a new type of GalNAc alpha 2,6-sialyltransferase (ST6GalNAc II) with a structure similar to that of a previously cloned GalNAc alpha 2,6-sialyltransferase (ST6GalNAc I; Kurosawa, N., Hamamoto, T., Lee, Y.-C., Nakaoka, T., Kojima, N., and Tsuji, S. (1994) J. Biol. Chem. 269, 1402-1409) was obtained from chicken testes. The predicted amino acid sequence of ST6GalNAc II encodes a protein with type II transmembrane topology, as found for other glycosyltransferases, and showed 32% identity with that of ST6GalNAc I. Transfection of the full length ST6GalNAc II gene into COS cells led to GalNAc alpha 2,6-sialyltransferase activity with a different substrate specificity from that of ST6GalNAc I. Moreover, asialofetuin after treatment with beta-galactosidase did not serve as an acceptor for this enzyme. 14C-Sialylated oligosaccharides obtained from resialylated asialobovine submaxillary mucin with this enzyme were identical to Gal beta 1,3([14C]NeuAc alpha 2,6)GalNAc-ol but not [14C]NeuAc alpha 2,6GalNAc-ol. These results clearly show that the expressed enzyme is a novel type of sialyltransferase that requires beta-galactoside residues linked to GalNAc residues, whereas sialic acid residues linked to galactose residues are not essential for the activity.
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PMID:Cloning and expression of Gal beta 1,3GalNAc-specific GalNAc alpha 2,6-sialyltransferase. 803 63

This work describes a lambda phage expression system, lambda foo, that produces foreign proteins fused to the surface of the virus particle. The lambda foo vector has multiple cloning sites for the insertion of a foreign DNA fragment and color selection for recombinants. Foreign proteins are fused to the C terminus of a truncated phage tail protein, pV, by a peptide linker. Conditional chain termination allows the assembly and fusion of multisubunit proteins. We have attached the complete Escherichia coli beta-galactosidase and the plant Bauhinia purpurea agglutinin by cloning their genes into the vector. The constructs express functionally active proteins on the phage particle surface and have been purified by affinity chromatography with an antibody for beta-galactosidase and a mucin as a ligand for Bauhinia purpurea agglutinin.
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PMID:Lambda foo: a lambda phage vector for the expression of foreign proteins. 805 94

The target molecules on the cell surface of Trypanosoma cruzi trypomastigotes reacting with lytic anti-alpha-galactosyl antibodies from chronic patients with Chagas' disease (Ch anti-Gal) have been purified by solvent extraction and identified as glycoconjugates migrating in the 74-96-kDa range (F2 antigen) and in the 120-200-kDa range (F3 antigen) on SDS-PAGE. The F3 antigen was tested for binding to Ch and normal human serum (NHS) anti-Gal and to MoAb 3C9. We observed that Ch anti-Gal and MoAb 3C9, but not NHS anti-Gal, bind strongly to the trypomastigote glycoconjugates. These antibodies, however, did not compete with each other for binding to F3 molecules, indicating that they are recognizing different epitopes. Binding of Ch anti-Gal to F3 antigen is abolished by treatment of these molecules with alpha- but not beta-galactosidase. Binding of 3C9 MoAb is abolished by treatment of F3 with sialidase. F2/F3 antigens absorbed Ch anti-Gal as well as lytic antibodies from total chagasic sera. These antigens also specifically discriminate between the serum reactivity of patients with active Chagas' disease and those of sera from cured patients, drug-treated patients with dissociated serology (positive conventional serology, negative trypanolytic activity), healthy individuals, and patients with several other infectious diseases. We also observed that F2/F3 antigens are anchored to the parasite membrane via glycosylphosphatidylinositol (GPI). The alpha-galactosyl epitopes recognized by Ch anti-Gal are present in a series of O-linked oligosaccharide chains in the mucin-like glycoprotein component of the complex.
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PMID:GPI-anchored glycoconjugates from Trypanosoma cruzi trypomastigotes are recognized by lytic anti-alpha-galactosyl antibodies isolated from patients with chronic Chagas' disease. 808 Dec 63

The purpose of the present paper was to study the effects of a dietary undigestible carbohydrate and intestinal microflora on mucin distribution (neutral, acid, sulphonated), glycolytic activities: beta-D-galactosidase (EC 3.2.1.23), N-acetyl-beta-D-galactosaminidase (EC 3.2.1.43), N-acetyl-beta-D-glucosaminidase (EC 3.2.1.30), alpha-L-fucosidase (EC 3.2.1.51) and bacterial metabolism (gas production, short-chain fatty acids (SCFA) and lactic acid caecal concentration) in germ-free (GF), conventional (CV) and heteroxenic (HE) rats (GF rats associated with a human flora). Rats were fed on either a control diet or a diet containing 40 g trans-galactosylated oligosaccharide (TOS)/kg. In GF rats fed on the control diet caecal pH was almost neutral and glycolytic activities negligible. The number of mucus-containing cells increased from the caecum to the colon for the three types of mucin. TOS had no effect in the caecum but it modified mucin cell repartition in the colon. In CV and HE rats fed on the control diet caecal pH was similar (6.8), but caecal SCFA and lactic acid concentrations (mumol/g) and gas production (ml/24 h) were higher in CV (70, 5.9 and 2.3 respectively) than in HE rats (32, 4.6 and 0.4 respectively). In CV, as in HE rats, acid-mucin-containing cells increased from the caecum to the colon and glycolytic activities were similar. TOS reduced acid-mucin-containing cells in the caecum of CV rats by twofold but had no effect in either the caecum or the colon of HE rats. TOS strongly increased beta-galactosidase activity and slightly modified the other glycolytic activities. Its effect on bacterial metabolites depended on bacterial status. However, comparison between CV and HE rats showed no evident relationship between the number of mucus-containing cells and measured bacterial metabolites. Differences between CV and HE rats might be due to bacterial microflora specificity. TOS had an intrinsic effect on mucus cell distribution in the colon of GF rats. In CV and HE rats the presence of the flora abolished this effect.
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PMID:Effects of galacto-oligosaccharide and bacterial status on mucin distribution in mucosa and on large intestine fermentation in rats. 832 64

A novel sulphotransferase (sulpho-T) activity from rat colonic mucosa was characterized using O-glycan core 1 substrate, Gal beta 1-3GalNAc alpha-benzyl. Derivatives of Gal beta 1-3GalNAc- were used to demonstrate that the 3- and 4-hydroxyl of Gal and the 2-acetamido group of the GalNAc residue of Gal beta 1-3GalNAc alpha-benzyl substrates were important for activity. Sulphated product using Gal beta 1-3GalNAc alpha-benzyl as substrate was analysed by ion spray mass spectrometry, methylation analysis, high-pH anion-exchange chromatography and beta-galactosidase digestion. The results suggested that sulphate was added to the 3-position of the Gal residue. The synthesis of core 2 from core 1 by UDP-GlcNAc: Gal beta 1-3GalNAc beta 6-GlcNAc-transferase was inhibited by sulphation of the Gal residue, indicating that GlcNAc beta 1-6 branching has to precede sulphation in the O-glycan core 1 processing pathway. These data demonstrate several novel pathways in the synthesis of sulphated mucin-type oligosaccharides.
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PMID:Characterization of a novel mucin sulphotransferase activity synthesizing sulphated O-glycan core 1,3-sulphate-Gal beta 1-3GalNAc alpha-R. 860 71


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