Tord Holme

Structural studies on the O-specific side-chains of the cell-wall lipopolysaccharide from Salmonella typhimurium 395 ms

The structure of the O-specific side-chains of the cell-wall lipopolysaccharide of Salmonella typhimurium 395 MS has been investigated. Methylation analyses of the original lipopolysaccharide, of the material obtained on mild hydrolysis of the lipopolysaccharide with acid, and of a product obtained by acetalation of all of the, free hydroxyl groups in the lipopolysaccharide, followed by alkaline deacetylation, have provided the essential information in this study. The mixtures of sugars obtained on acid hydrolysis of the different methylated products were analysed, as alditol acetates, by g.l.c.-mass spectrometry. As a result of these studies, a detailed structure of the repeating unit of these side-chains is presented.

Structural studies of the Vibrio cholerae O-antigen.

The dominant part of the O-antigen of Vibrio cholerae is a homopolysaccharide composed of (1 leads to 2)-linked 4-amino-4,6-dideoxy-alpha-D-mannopyranosyl (perosaminyl) residues, in amino groups of which are acylated by 3-deoxy-L-glycero-tetronic acid. Most of the amino sugar is decomposed during acid hydrolysis. Treatment of the polymer with anhydrous hydrogen fluoride, which cleaves the glycosidic linkages but does not cause N-deacylation, followed by acid hydrolysis under mild conditions, produced the monomer in good yield. Treatment of the N-deacylated polysaccharide with nitrous acid caused deamination with concomitant rearrangements, typical of 4-amino-4-deoxyhexopyranosyl residues in which the amino group occupies an equatorial position.

Studies on extracellular proteolytic enzymes from Staphylococcus aureus: I. Purification and characterization of one neutral and one alkaline protease

Abstract Two extracellular proteolytic enzymes from Staphylococcus aureus, strain V8, were separated by isoelectric focusing. One protease, with an isoelectric point at pH 4.0 (protease I) was purified 185-fold by negative adsorption on CM-Sephadex, isoelectric focusing and gel chromatography on Sephadex G-75. The molecular weight of this enzyme was about 21 000. The pH-optimum was at 7.8 using casein as the substrate, and at 5.0 and 7.0–9.0 using hemoglobin as the substrate. The activity of protease I was not significantly affected by divalent cations, metal-chelating agents, sulfhydryl reagents, or by diisopropylfluorophosphate. The other protease with an isoelectric point at 9.4 (protease II) was purified 310-fold by negative adsorption on DEAE-Sephadex, isoelectric focusing and gel chromatography on BioGel P 60. The molecular weight was 12 500. The pH optimum was at pH 8.8. Protease II was only active in the presence of reducing agents, and was inactivated by heavy metals. Protease II exhibited esterase activity, using N- benzoyl - l - tyrosine ethyl ester as the substrate.

Structural studies of the O-polysaccharide from the lipopolysaccharide of Moraxella (Branhamella) catarrhalis serotype A (strain ATCC 25238).

The polysaccharide of the Moraxella (Branhamella) catarrhalis serotype A lipopolysaccharide was prepared by mild acid hydrolysis followed by gel permeation chromatography. The structure was established by methylation analysis, mass spectrometry, and NMR spectroscopy. It is concluded that the O-antigenic polysaccharide has the following structure. [formula see text] Methylation analysis of the intact lipopolysaccharide showed that the lipid A portion consisted of 6-substituted glucosamine residues. Methylation followed by methanolysis showed that two Kdo residues were present, one terminal and one 4,5-substituted residue. A terminal Kdo thus substitutes the branch-point Kdo in the 4-position.

Chemical Composition of Cell-wall Polysaccharide of Rough Mutants of Salmonella typhimurium

SUMMARY: Quantitative analyses of monosaccharide constituents of the cell-wall polysaccharide of the smooth form and rough mutants of Salmonella typhimurium were made by gas-liquid chromatography of glycitol acetates produced from acid hydrolysates of phenol+water extracts of cell-wall preparations. The presence, in varying amounts, of sugars characteristic of the S-specific repeating unit was detected in all rough mutants investigated. Definite conclusions about the core structure of the different rough mutants could not be drawn by using only the basis of the monosaccharide composition of the cell-wall polysaccharide. Ribose was found in the cell-wall polysaccharide of the smooth form in small amounts.

Structural studies of the O-antigen oligosaccharides from two strains of Moraxella catarrhalis serotype C

The oligosaccharide parts from Moraxella (Branhamella) catarrhalis serotype C lipooligosaccharides were isolated by mild acid hydrolysis followed by gel permeation chromatography. Four different oligosaccharides could be identified from strain RS26 and two from strain RS10. The structures of the O-oligosaccharides were established by methylation analyses, mass spectrometry, and NMR spectroscopy. It is concluded that the oligosaccharide O-antigens from RS26 are a mixture of octa-, deca-, and undeca-saccharides, and most likely a heptasaccharide. Strain RS10 contains the deca- and the undeca-saccharide only. The structures for the oligosaccharides are shown below. [formula: see text] OS(7) [formula: see text] OS(8) [formula: see text] OS(10) [formula: see text] OS(11) Methylation analysis of the intact lipooligosaccharides showed that two Kdo residues were present, one terminal and one 4,5-substituted residue. It also showed that they consisted of a lipid A portion with 6-substituted glucosamine residues.

The structures of oligosaccharides isolated from the lipopolysaccharide of Moraxella catarrhalis serotype B, strain CCUG 3292

The oligosaccharides from the lipopolysaccharides of Moraxella catarrhalis serotype B, strain CCUG 3292, were isolated after mild acid hydrolysis and separated by high-performance anion-exchange chromatography. The structures of the oligosaccharides were established by fast atom bombardment mass spectrometry and nuclear magnetic resonance spectroscopy. It is concluded that the oligosaccharides comprise a mixture of mainly a nona- and a deca-saccharide. [formula: see text] Smaller amounts of undeca-saccharides and of truncated forms, namely, hexa-, hepta-, and octa-saccharides, were also detected.

Structural studies of the O-antigen from Vibrio cholerae O:21

The O-antigen from Vibrio cholerae O:21 has been investigated, using n.m.r. spectroscopy, methylation analysis, and Smith degradation as the main methods. It is concluded that the O-antigen is composed of tetrasaccharide repeating-units having the following structure (in which Hep = D-glycero-D-manno-heptose). (Formula: see text).

Lack of serotype-specific antibody response to lipopolysaccharide antigens ofMoraxella catarrhalis during lower respiratory tract infection

An enzyme immunoassay (EIA) was used to determine the antibody response to different serotypes of lipopolysaccharide (LPS) antigens ofMoraxella catarrhalis in adult patients with lower respiratory tract infections (LRTI).Moraxella catarrhalis was isolated from sputum or nasopharyngeal samples from 20 patients with LRTI. Sixteen of the isolates were serotype A, four were type B and none were type C. The antibody response to the different LPS serotypes was determined in paired sera from patients suffering from LRTI. In addition to the 20 patients withMoraxella catarrhalis isolated (Group 1), a group of seven patients with LRTI of unknown etiology (Group 2) and a group of ten patients with LRTI of known other bacterial etiology (Group 3) were selected for this study. An increase in antibody levels of > 1.5-fold (convalescent/acute-phase serum) was recorded in approximately half of the patients, not only in the first group (Moraxella catarrhalis isolated) but also in the other two groups. However, in the first and second groups there was a correlation between an increase in antibody levels in the LPS EIA and in an EIA using whole bacterial cells as antigen. In the group of patients in whomMoraxella catarrhalis was isolated, the antibody response to LPS antigens was not serotype specific. The antibody response to type-A and type-B LPS was more predominant than the response to type-C LPS in most patients. The results show that an antibody response to LPS antigens during lower respiratory tract infections caused byMoraxella catarrhalis could be demonstrated and that there was a correlation between the response found in a whole-cell EIA and the LPS EIA. However, titre increases, especially to type-A LPS, were recorded in seven of ten patients with LRTI of other bacterial etiology. Serological investigations based on determination of antibodies to LPS antigens were not useful for etiological studies but may be of interest for studying the pathogenetic mechanisms of respiratory tract infections.

The Inhibitory Effect of Coli-Endotoxin on Alpha-Adrenergic Receptor Functions in the Lower Urinary Tract: An In vitro Study in Cats

The effects of endotoxin from E. coli 06 on alpha-adrenergic receptor functions have been investigated in muscle strips from the outlet region of the feline bladder. The endotoxin from this strain, known to cause urinary tract infections, inhibited the alpha-adrenergic contractions induced by phenylephrine and noradrenaline. This inhibition was dose-dependent, correlated to time of exposure, and reversible. Alpha-adrenergic receptor function in muscle strips from cat aorta was also inhibited, but not cholinergic stimulation of muscle segments from the bladder.