Preparation and structural analysis of oligosaccharide monophosphates obtained from the lipopolysaccharide of recombinant strains of Salmonella minnesota and Escherichia coli expressing the genus-specific epitope of Chlamydia lipopolysaccharide.
The lipopolysaccharide of the recombinant strain Salmonella minnesota r595-207 expressing the genus-specific epitope of Chlamydia lipopolysaccharide [Holst, O., Brade, L., Kosma, P. and Brade, H. (1991) J. Bacteriol, 173, 1862-1866] was sequentially de-O- and de-N-acylated by mild hydrazinolysis and treatment with 4 M KOH, respectively. The resulting mixture of compounds was separated by high-performance anion-exchange chromatography and gel-permeation chromatography, yielding four oligosaccharide phosphates two of which were readily identified by their 1H-NMR- and 13C-NMR spectra as alpha-Kdo-(2-4)-alpha-Kdo-(2-6)-beta-D-GlcpN-(1-6)-alpha-D-Glcp N 1,4'-bisphosphate (tetrasaccharide bisphosphate; Kdo = 3-deoxy-D-manno-octulopyranosonic acid) and alpha-Kdo-(2-8)-alpha-Kdo-(2-4)-alpha-Kdo-(2-6)-beta-D-GlcpN-(1-6) -alpha-D- GlcpN 1,4'-bisphosphate (pentasaccharide bisphosphate) [Holst, O., Broer, W., Thomas-Oates, J.E., Mamat, U. and Brade, H. (1993) Eur. J. Biochem. 214, 703-710]. The structures of the other two compounds were established by chemical analysis, NMR spectroscopy, and fast-atom-bombardment mass spectrometry as alpha-Kdo- (2-4)-alpha-Kdo-(2-6)-beta-D-GlcpN-(1-6)-alpha-D-GlcpN 1-phosphate (tetrasaccharide 1-phosphate) and alpha-Kdo-(2-8)-alpha-Kdo-(2-4)-alpha-Kdo-(2-6)-beta-D-GlcpN-(1-6) -alpha-D- GlcpN 1-phosphate (pentasaccharide 1-phosphate). alpha-Kdo-(2-4)-alpha-Kdo-(2-6)-beta-D-GlcpN-(1-6)-alpha/beta- D-GlcpN 4'-phosphate (tetrasaccharide 4'-phosphate) and alpha-Kdo-(2-8)-alpha-Kdo-(2-4)-alpha-Kdo-(2-6)-beta-D-GlcpN-(1-6) -alpha/beta-D-GlcpN 4'-phosphate (pentasaccharide 4'-phosphate) were prepared from the 1,4'-bisphosphates isolated from the recombinant strain Escherichia coli F515-207 by treatment with alkaline phosphatase and purification by high-performance anion-exchange chromatography and gel-permeation chromatography. Their structures were characterised by chemical analysis, NMR spectroscopy, and fast-bombardment mass spectrometry.
Holst
O
O
Division of Biochemical Microbiology, Forschungsinstitut Borstel, Germany.
Thomas-Oates
J E
JE
Brade
H
H
D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D002689:Chlamydia / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D000939:Epitopes / Q000235:genetics; D004926:Escherichia coli / Q000737:chemistry* / Q000235:genetics; D008070:Lipopolysaccharides / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000032:analysis / Q000737:chemistry*; D009943:Organophosphorus Compounds / Q000032:analysis / Q000737:chemistry*; D011995:Recombination, Genetic; D012475:Salmonella / Q000737:chemistry* / Q000235:genetics; D016339:Spectrometry, Mass, Fast Atom Bombardment
D000939:Epitopes; D008070:Lipopolysaccharides; D009844:Oligosaccharides; D009943:Organophosphorus Compounds