scholarly journals Purification and characterization of CS2, a sialic acid-specific haemagglutinin of enterotoxigenic Escherichia coli

1988 ◽  
Vol 255 (1) ◽  
pp. 105-111 ◽  
Author(s):  
P O Sjöberg ◽  
M Lindahl ◽  
J Porath ◽  
T Wadström
Keyword(s):  
Escherichia Coli ◽  
Sialic Acid ◽  
Gel Filtration ◽  
Enterotoxigenic Escherichia Coli ◽  
Guanidinium Chloride ◽  
Neuraminidase Treatment ◽  
Low Concentrations ◽  
Sepharose 4B ◽  
Bovine Erythrocytes

CS2 fimbriae of enterotoxigenic Escherichia coli were purified and characterized. The surface haemagglutinins (fimbriae) were detached by sonication from a strain producing only the CS2 fimbriae. Isolation was carried out by gel filtration on a Sepharose 4B column. After depolymerization, the fimbriae subunits were purified on a Sephacryl S-300 column in 8.0 M-guanidinium chloride. From 1 litre of medium, 4-6 mg of purified fimbriae was obtained. We found that CS2 fimbriae were completely dissociated by saturated guanidinium chloride into subunits with a molecular mass of 16.5 kDa. CS2 fimbriae was sialic acid-specific, since sialic acids were the most potent inhibitors, and neuraminidase treatment of erythrocytes abolished haemagglutination. Both fimbriae and fimbrial subunits were found to bind to bovine erythrocytes. The binding of subunits to erythrocytes could be inhibited with low concentrations of sialyl-lactose.

scholarly journals Studies on gastric mucoproteins. The isolation and characterization of the mucoprotein of the water-soluble mucus from pig cardiac gastric mucosa

1971 ◽  
Vol 123 (5) ◽  
pp. 845-853 ◽  
Author(s):  
D. Snary ◽  
A. Allen
Keyword(s):  
Sialic Acid ◽  
Gastric Mucosa ◽  
Gel Filtration ◽  
Water Soluble ◽  
Group Activities ◽  
Isolation And Characterization ◽  
Two Peaks ◽  
Sepharose 4B

1. Gel filtration of the water-soluble radioactive mucus produced three radioactive fractions, fraction A excluded on Sepharose 4B, fraction B included on Sepharose 4B but excluded on Sephadex G-200, and fraction C included on Sephadex G-200. 2. The specific radioactivities of fractions A and B were the same, with fraction C a little lower, whether the material was labelled with 14C-labelled carbohydrate or with 3H-labelled protein prepared by incubation of mucosal scrapings in vitro with [U-14C]glucose or [G-3H]threonine respectively. 3. Fractions A and B had an analysis of protein 22%, hexose 28%, hexosamine 28%, fucose 10% and sialic acid 1%; fraction C had an analysis closely similar to this, except that it contained about 10% of a protein contaminant. 4. All three fractions had closely similar A and H blood-group activities. 5. Ultracentrifuge studies showed fractions A, B and C were polydisperse with s025,w values of 18.7S, 4.9S and 3.9S respectively. 6. The unfractionated water-soluble mucus contained only two peaks, fraction A 18.7S and a peak of 4.4S, which was a combination of fractions B and C. 7. The radioactive mucoprotein accounted for 85% by weight of the soluble mucus and the results show that it consisted of two distinct fractions A and B–C, which were chemically, biosynthetically and immunologically very similar.

scholarly journals Effects of pronase and neuraminidase treatment on a myelin-associated glycoprotein in developing brain

1976 ◽  
Vol 156 (1) ◽  
pp. 143-150 ◽  
Author(s):  
R H Quarles
Keyword(s):  
Molecular Weight ◽  
Sialic Acid ◽  
Sodium Dodecyl Sulphate ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Apparent Molecular Weight ◽  
Weight Class ◽  
Developmental Difference ◽  
Adult Rats ◽  
Neuraminidase Treatment

Rats (14 days old) were injected with [14c]fucose and young adult rats with [3H]fucose in order to label the myelin-associated glycoproteins. As previously reported, the major [14C]fucose-labelled glycoprotein in the immature myelin had a higher apparent molecular weight on sodium dodecyl sulphate/polyacrylamide gels that the [3H]fucose-labelled glycoprotein in mature myelin. This predominant doubly labelled glycoprotein component was partially purified by preparative gel electrophoresis and converted to glycopeptides by extensive Pronase digestion. Gel filtration on Sephadex G-50 separated the glycopeptides into several clases, which were designted A,B, C AND D, from high to low molecular weight. The 14C-labelled glycopeptides from immature myeline were enriched in the highest-molecular-weight class A relative to the 3H-labelled glycopeptides from mature myelin. Neuraminidase treatment of the glycoprotein before Pronase digestion greatly decreased the proportion of glycopeptides fractionating in the higher-molecular-weight classes and largely eliminated the developmental differences that were apparent by gel filtration. However, neuraminidase treatment did not decrease the magnitude of the developmental difference revealed by electrophoresing the intact glycoprotein on sodium dodecyl sulphate gels, although it did decrease the apparent molecular weight of the glycoprotein from both the 15-day-old and adult rats by an amount comparable in magnitude to that developmental difference. The results from gel filtration of glycopeptides indicate that there is a higher content of large molecular weight, sialic acid-rich oligosaccharide units in the glycoprotein of immature myelin. However, the higher apparent molecular weight for the glycoprotein from 15-day-old rats on sodium dodcyl sulphate gels is not due primarily to its higher sialic acid content.

scholarly journals Occurrence of adhesins causing mannose-resistant haemagglutination of bovine erythrocytes in enterotoxigenic Escherichia coli

1979 ◽  
Vol 5 (2) ◽  
pp. 85-90 ◽  
Author(s):  
C.J. Smyth ◽  
B. Kaijser ◽  
E. Bäck ◽  
A. Faris ◽  
R. Möllby ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Enterotoxigenic Escherichia Coli ◽  
Bovine Erythrocytes

scholarly journals Characterization of the F41 fimbrial antigen of enterotoxigenic Escherichia coli by using monoclonal antibodies.

1989 ◽  
Vol 57 (4) ◽  
pp. 1192-1199 ◽  
Author(s):  
F G van Zijderveld ◽  
F Westenbrink ◽  
J Anakotta ◽  
R A Brouwers ◽  
A M van Zijderveld
Keyword(s):  
Escherichia Coli ◽  
Monoclonal Antibodies ◽  
Enterotoxigenic Escherichia Coli

scholarly journals Purification and Characterization of the Recombinant Thermus sp. Strain T2 α-Galactosidase Expressed in Escherichia coli

2001 ◽  
Vol 67 (4) ◽  
pp. 1601-1606 ◽  
Author(s):  
Mitsunori Ishiguro ◽  
Satoshi Kaneko ◽  
Atsushi Kuno ◽  
Yoshinori Koyama ◽  
Shigeki Yoshida ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Polyacrylamide Gel Electrophoresis ◽  
Active Form ◽  
Gel Filtration ◽  
Amino Acid Sequences ◽  
Side Chain ◽  
Catalytic Function ◽  
Native Polyacrylamide Gel Electrophoresis

ABSTRACT The nucleotide sequence of the Thermus sp. strain T2 DNA coding for a thermostable α-galactosidase was determined. The deduced amino acid sequence of the enzyme predicts a polypeptide of 474 amino acids (M r, 53,514). The observed homology between the deduced amino acid sequences of the enzyme and α-galactosidase from Thermus brockianus was over 70%.Thermus sp. strain T2 α-galactosidase was expressed in its active form in Escherichia coli and purified. Native polyacrylamide gel electrophoresis and gel filtration chromatography data suggest that the enzyme is octameric. The enzyme was most active at 75°C forp-nitrophenyl-α-d-galactopyranoside hydrolysis, and it retained 50% of its initial activity after 1 h of incubation at 70°C. The enzyme was extremely stable over a broad range of pH (pH 6 to 13) after treatment at 40°C for 1 h. The enzyme acted on the terminal α-galactosyl residue, not on the side chain residue, of the galactomanno-oligosaccharides as well as those of yeasts and Mortierella vinacea α-galactosidase I. The enzyme has only one Cys residue in the molecule.para-Chloromercuribenzoic acid completely inhibited the enzyme but did not affect the mutant enzyme which contained Ala instead of Cys, indicating that this Cys residue is not responsible for its catalytic function.

Chemical characterization of urinary glycopeptides.

1987 ◽  
Vol 33 (12) ◽  
pp. 2214-2219
Author(s):  
H Matsue ◽  
K Takagaki ◽  
K Honda ◽  
Y Nakagawa ◽  
F Gejyo ◽  
...  
Keyword(s):  
Sialic Acid ◽  
Carbohydrate Chain ◽  
Cetylpyridinium Chloride ◽  
Gel Filtration ◽  
Chemical Characterization ◽  
Small Peptide ◽  
Supernatant Liquid ◽  
Peptide Moiety ◽  
Peptide Linkage

Abstract We prepared human urinary glycopeptides from the supernatant liquid remaining after precipitation of the nondialyzable fraction with cetylpyridinium chloride. Using cation exchange and affinity chromatographies and gel filtration, we obtained 28 glycopeptide subfractions. By compositional analyses of sugar and amino acid, and by reducing-terminal analyses after reduction with NaBH4, we determined the size of the carbohydrate moiety and the types of carbohydrate-peptide linkage involved. We isolated several glycopeptides not previously described: six with sialic acid, two with fucose, two with glucose, and one with N-acetylgalactosamine. The sialic acid glycopeptides had a short carbohydrate chain of the O-glycoside type. The fucose-containing glycopeptides were fucosyllactosaminoglycans. The glucose glycopeptides were polymers linked to a small peptide moiety. The N-acetylgalactosamine-rich glycopeptide was found in an N-glycoside-type fraction, with N-acetylgalactosamine at the nonreducing terminal.

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