Structural studies of some bacterial polysaccharides and extension of a method for lipid A cleavage

University dissertation from Stockholm : Karolinska Institutet, Clinical Research Center

Abstract: The focus of the work presented in this thesis has been to study the structures of the capsular polysaccharide from Serratia marcescens O14:K12, and the O-antigen polysaccharides from Hafnia alvei strain 10457 and Vibrio cholerae serogroup 037. Nuclear magnetic resonance (NMR) spectroscopy was the primary technique employed. S. marcescens O14:K12 contains both an O-antigen polysaccharide and a capsular polysaccharide (K-antigen). The K-antigen was successfully purified from the mixture of the two polysaccharides and was analysed. The structure of the K-antigen is similar, but not identical, to other K-antigens of S. marcescens In addition, the polysaccharide has two acidic components, a pyruvic acid and a uronic acid, which is unusual. The study is a step towards the identification of the serology of S. marcescens. New types of Hafnia alvei strains from Bangladesh have been reported to cause severe diarrhoea. However, these strains also contain a virulence factor that is characteristic for enteropathogenic Escherichia coli (EPEC) making the serologic classification uncertain. The Hafnia strains from Bangladesh may be pathogenic H. alvei, unusual biotypes of E. coli, or represent a new species in the genus Escherichia. From the data on the structure of the 0antigen, it may be classified as either of the two with regard to carbohydrate structure. An unusual feature, however, is the terminally linked N-acetyl neuraminic acid, which is a common constituent in glycoproteins and glycolipids. Vibrio cholerae serogroup 037 has genetic relationships with the cholera causing V. cholerae serogroups 01 and 0139. This prompted us to investigate the O-antigen structure from V. cholerae 037 Sudan strain. The structure of the repeating unit was established, which differed significantly from 01 and 0139. The last part of this thesis describes an improved method for cleaving off lipid A from the LPS. The procedure was shown to work on LPS from.S. marcescens O14:K12, H. alvei strain 10457, and E. coli 0128. This method involves ceric ammonium nitrate (CAN) in water and may be a useful alternative if polysaccharides from LPS contain acid-labile constituents that might be cleaved off during normal delipidation conditions.

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