scholarly journals A New Look at the Enterobacterial Common Antigen Forms Obtained during Rough Lipopolysaccharides Purification

2021 ◽  
Vol 22 (2) ◽  
pp. 701
Author(s):  
Tomasz K Gozdziewicz ◽  
Anna Maciejewska ◽  
Alona Tsybulska ◽  
Czeslaw Lugowski ◽  
Jolanta Lukasiewicz
Keyword(s):  
Current Knowledge ◽  
Membrane Integrity ◽  
Gel Filtration ◽  
Common Antigen ◽  
Mild Acid Hydrolysis ◽  
Enterobacterial Common Antigen ◽  
Lipopolysaccharide Endotoxin ◽  
Lipid Moiety ◽  
Detailed Composition ◽  
Mild Acid

Enterobacterial common antigen (ECA) is a conserved antigen expressed by enterobacteria. It is built by trisaccharide repeating units: →3)-α-D-Fucp4NAc-(1→4)-β-D-ManpNAcA-(1→4)-α-D-GlcpNAc-(1→ and occurs in three forms: as surface-bound linear polysaccharides linked to a phosphoglyceride (ECAPG) or lipopolysaccharide − endotoxin (ECALPS), and cyclic form (ECACYC). ECA maintains, outer membrane integrity, immunogenicity, and viability of enterobacteria. A supernatant obtained after LPS ultracentrifugation was reported as a source for ECA isolation, but it has never been assessed for detailed composition besides ECACYC. We used mild acid hydrolysis and gel filtration, or zwitterionic-hydrophilic interaction liquid (ZIC®HILIC) chromatography combined with mass spectrometry for purification, fractionation, and structural analysis of rough Shigella sonnei and Escherichia coli R1 and K12 crude LPS preparations. Presented work is the first report concerning complex characteristic of all ECA forms present in LPS-derived supernatants. We demonstrated high heterogeneity of the supernatant-derived ECA that contaminate LPS purified by ultracentrifugation. Not only previously reported O-acetylated tetrameric, pentameric, and hexameric ECACYC have been identified, but also devoid of lipid moiety linear ECA built from 7 to 11 repeating units. Described results were common for all selected strains. The origin of linear ECA is discussed against the current knowledge about ECAPG and ECALPS.

scholarly journals Studies of the polysaccharide fraction from the lipopolysaccharide of Pseudomonas alcaligenes

1974 ◽  
Vol 139 (3) ◽  
pp. 633-643 ◽  
Author(s):  
James A. Lomax ◽  
George W. Gray ◽  
Stephen G. Wilkinson
Keyword(s):  
Gel Filtration ◽  
Periodate Oxidation ◽  
Side Chain ◽  
Partial Hydrolysis ◽  
Methylation Analysis ◽  
Mild Acid Hydrolysis ◽  
Pseudomonas Alcaligenes ◽  
Crude Polysaccharide ◽  
Hydrolysis Of ◽  
Mild Acid

Studies of the lipopolysaccharide of Pseudomonas alcaligenes strain BR 1/2 were extended to the polysaccharide moiety. The crude polysaccharide, obtained by mild acid hydrolysis of the lipopolysaccharide, was fractionated by gel filtration. The major fraction was the phosphorylated polysaccharide, for which the approximate proportions of residues were; glucose (2), rhamnose (0.7), heptose (2–3), galactosamine (1), alanine (1), 3-deoxy-2-octulonic acid (1), phosphorus (5–6). The heptose was l-glycero-d-manno-heptose. The minor fractions from gel filtration contained free 3-deoxy-2-octulonic acid, Pi and PPi. The purified polysaccharide was studied by periodate oxidation, methylation analysis, partial hydrolysis, and dephosphorylation. All the rhamnose and part of the glucose and heptose occur as non-reducing terminal residues. Other glucose residues are 3-substituted, and most heptose residues are esterified with condensed phosphate residues, possibly in the C-4 position. Free heptose and a heptosylglucose were isolated from a partial hydrolysate of the polysaccharide. The location of galactosamine in the polysaccharide was not established, but either the C-3 or C-4 position appears to be substituted and a linkage to alanine was indicated. In its composition, the polysaccharide from Ps. alcaligenes resembles core polysaccharides from other pseudomonads: no possible side-chain polysaccharide was detected.

scholarly journals SYNTHESIS OF GLYCOPROTEINS IN A SINGLE IDENTIFIED NEURON OF APLYSIA CALIFORNICA

1974 ◽  
Vol 61 (3) ◽  
pp. 649-664 ◽  
Author(s):  
Richard T. Ambron ◽  
James E. Goldman ◽  
Elizabeth Barnes Thompson ◽  
James H. Schwartz
Keyword(s):  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Apparent Molecular Weight ◽  
Mild Acid Hydrolysis ◽  
Giant Neuron ◽  
Electrophoretic Patterns ◽  
Identified Neuron ◽  
Hydrolysis Of ◽  
Mild Acid

Incorporation of L-[3H]fucose into glycoproteins was studied in R2, the giant neuron in the abdominal ganglion of Aplysia. [3H]fucose injected directly into the cell body of R2 was readily incorporated into glycoproteins which, as shown by autoradiography, were confined almost entirely to the injected neuron. Within 4 h after injection, 67% of the radioactivity in R2 had been incorporated into glycoproteins; at least 95% of these could be sedimented by centrifugation at 105,000 g, suggesting that they are associated with membranes. Extraction of the particulate fraction with sodium dodecyl sulfate (SDS), followed by gel filtration on Sephadex G-200 and polyacrylamide gel electrophoresis in SDS revealed the presence of only five major radioactive glycoprotein components which ranged in apparent molecular weight from 100,000 to 200,000 daltons. Similar results were obtained after intrasomatic injection of [3H]N-acetylgalactosamine. Mild acid hydrolysis of particulate fractions released all of the radioactivity in the form of fucose. When ganglia were incubated in the presence of [3H]fucose, radioactivity was preferentially incorporated into glial cells and connective tissue. In contrast to the relatively simple electrophoretic patterns obtained from cells injected with [3H]fucose, gel profiles of particulate fractions labeled with [14C]valine were much more complex.

scholarly journals An “Uncommon” Role for Cyclic Enterobacterial Common Antigen in Maintaining Outer Membrane Integrity

mBio ◽  
2018 ◽  
Vol 9 (5) ◽  
Author(s):  
Corey S. Westfall
Keyword(s):  
Membrane Protein ◽  
Outer Membrane ◽  
Membrane Integrity ◽  
Physiological Role ◽  
Permeability Barrier ◽  
Common Antigen ◽  
Enterobacterial Common Antigen ◽  
Inner Membrane Protein ◽  
Outer Membrane Permeability

ABSTRACTAlthough discovered over 50 years ago, the physiological role of enterobacterial common antigen, a surface antigen produced by all members of theEnterobacteriaceae, has been poorly understood. In the work of Mitchell et al. (mBio 9:e01321-18, 2018,https://doi.org/10.1128/mBio.01321-18), the cyclized version of enterobacterial common antigen has been shown to play a role in maintaining the outer membrane permeability barrier, possibly through the inner membrane protein YhdP. This work also provides the tests needed to separate true effects from the numerous possible artifacts possible with mutations in enterobacterial common antigen synthesis.

scholarly journals Bi- and tri-antennary human transferrin glycopeptides and their affinities for the hepatic lectin specific for asialo-glycoproteins

1979 ◽  
Vol 181 (3) ◽  
pp. 633-638 ◽  
Author(s):  
Mark W. C. Hatton ◽  
Leopold März ◽  
Leslie R. Berry ◽  
Maria T. Debanne ◽  
Erwin Regoeczi
Keyword(s):  
Concanavalin A ◽  
Gel Filtration ◽  
Structural Heterogeneity ◽  
Anion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Galactose Oxidase ◽  
Molecular Forms ◽  
Mild Acid Hydrolysis ◽  
Human Transferrin ◽  
Mild Acid

Glycopeptides were isolated from a proteolytic digest of human transferrin. After mild acid hydrolysis the desialylated glycopeptides were labelled by the galactose oxidase/NaB3H4 procedure and then fractionated by Sephadex-gel filtration or by anion-exchange chromatography. Either technique allowed separation of the two heterosaccharide chains (designated glycan I and glycan II) previously described for this protein by Spik, Vandersyppe, Fournet, Bayard, Charet, Bouquelet, Strecker & Montreuil (1974) (in Actes du Colloque Internationale No. 221 vol. 1, pp. 483–499). Subsequent chromatography on Sepharose–concanavalin A separated fractions containing different quantities of carbohydrates for each glycan, as indicated by analyses. The isolated glycan fractions were then tested for their abilities to bind to the immobilized rabbit hepatic lectin. Our studies suggest that either glycan can have a bi- or tri-antennary structure. Desialylated biantennary glycans I and II did not bind to the hepatic lectin. Desialylated triantennary glycan I was slightly retarded by the hepatic lectin, whereas the triantennary glycan II consisted of equal quantities of a retarded and a bound type. Desialylated triantennary glycan II was totally displaced from the hepatic lectin by using a buffer containing 0.05m-EDTA. The results suggest that greater structural heterogeneity exists in the carbohydrate moiety of human transferrin than was previously envisaged. Such heterogeneity could be reflected in several molecular forms of human transferrin, which, after desialylation, differ significantly in their affinities for the hepatic lectin.

Amino acid sequence of cyanogen bromide fragment CB5 of human hemopexin

1989 ◽  
Vol 54 (3) ◽  
pp. 803-810 ◽  
Author(s):  
Ivan Kluh ◽  
Ladislav Morávek ◽  
Manfred Pavlík
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Acid Hydrolysis ◽  
Cyanogen Bromide ◽  
Polypeptide Chain ◽  
Amino Acid Residues ◽  
Mild Acid Hydrolysis ◽  
Methionine Residue ◽  
Cyanogen Bromide Fragment ◽  
Mild Acid

Cyanogen bromide fragment CB5 represents the region of the polypeptide chain of hemopexin between the fourth and fifth methionine residue (residues 232-352). It contains 120 amino acid residues in the following sequence: Arg-Cys-Ser-Pro-His-Leu-Val-Leu-Ser-Ala-Leu-Thr-Ser-Asp-Asn-His-Gly-Ala-Thr-Tyr-Ala-Phe-Ser-Gly-Thr-His-Tyr-Trp-Arg-Leu-Asp-Thr-Ser-Arg-Asp-Gly-Trp-His-Ser-Trp-Pro-Ile-Ala-His-Gln-Trp-Pro-Gln-Gly-Pro-Ser-Ala-Val-Asp-Ala-Ala-Phe-Ser-Trp-Glu-Glu-Lys-Leu-Tyr-Leu-Val-Gln-Gly-Thr-Gln-Val-Tyr-Val-Phe-Leu-Thr-Lys-Gly-Gly-Tyr-Thr-Leu-Val-Ser-Gly-Tyr-Pro-Lys-Arg-Leu-Glu-Lys-Glu-Val-Gly-Thr-Pro-His-Gly-Ile-Ile-Leu-Asp-Ser-Val-Asp-Ala-Ala-Phe-Ile-Cys-Pro-Gly-Ser-Ser-Arg-Leu-His-Ile-Met. The sequence was derived from the data on peptides prepared by cleavage of fragment CB5 by mild acid hydrolysis, by trypsin and chymotrypsin.

Pigments of marine animals. XI. Angular naphthopyrones from the crinoid Comanthus parvicirrus timorensis

1971 ◽  
Vol 24 (7) ◽  
pp. 1487 ◽  
Author(s):  
IR Smith ◽  
MD Sutherland
Keyword(s):  
Methyl Ether ◽  
Dry Weight ◽  
Water Soluble ◽  
Spectral Studies ◽  
Mild Acid Hydrolysis ◽  
Marine Animals ◽  
Yellow Colour ◽  
Methoxy Derivative ◽  
Colour Variant ◽  
Mild Acid

Green specimens of the comatulid crinoid, Comanthus parvicirrus timorensis J. Muller, yield to acetone three yellow water-soluble colouring matters, comaparvin sulphate, 6-methoxycomaparvin sulphate, and 6-methoxycomaparvin 5-methyl ether sulphate in approximately 0.1 %, 0.7 %, and 0.7 % yield respectively of the dry weight of the animal, Mild acid hydrolysis yields the corresponding phenols, the structures of which have been deduced largely by spectral studies as very probably 5,8-dihydroxy-10-methoxy-2-n-propyl-4H-naphtho[1,2-b]pyran-4-one (1), the 6-methoxy derivative of (1), and the 6-methoxy methyl ether of (1) respectively. A yellow colour variant of the same species yielded the same colouring matters in slightly different proportions. The calcareous skeleton contains what are probably polyhydroxynaphthoquinones in combined form.

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