Open AccessPreparation 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
Author(s) -
HOLST Otto,
THOMASOATES Jane E.,
BRADE Helmut
Publication year - 1994
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1994.tb18856.x
Subject(s) - tetrasaccharide , oligosaccharide , recombinant dna , nuclear magnetic resonance spectroscopy , chemistry , stereochemistry , escherichia coli , fast atom bombardment , carbohydrate conformation , alpha (finance) , biochemistry , mass spectrometry , chromatography , polysaccharide , medicine , construct validity , nursing , patient satisfaction , gene
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 1 H‐NMR‐ and 13 C‐NMR spectra as α‐Kdo‐(2–4)‐α‐Kdo‐(2–6)‐β‐D‐Glc p N‐(1–6)‐α‐D‐Glc p N 1,4′‐bisphosphate (tetrasaccharide bisphosphate; Kdo = 3‐deoxy‐D‐ manno ‐octulopyranosonic acid) and α‐Kdo‐(2–8)‐α‐Kdo‐(2–4)‐α‐Kdo‐(2–6)‐β‐D‐Glc p N‐(1–6)‐α‐D‐Glc p N 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 α‐Kdo‐(2–4)‐α‐Kdo‐(2–6)‐β‐D‐Glc p N‐(1–6)‐α‐D‐Glc p N 1‐phosphate (tetrasaccharide 1‐phosphate) and α‐Kdo‐(2–8)‐α‐Kdo‐(2–4)‐α‐Kdo‐(2–6)‐β‐D‐Glc p N‐(1–6)‐α‐D‐Glc p N 1‐phosphate (pentasaccharide 1‐phosphate). α‐Kdo‐(2–4)‐α‐Kdo‐(2–6)‐β‐D‐Glc p N‐(1–6)‐α/β‐D‐Glc p N 4′‐phosphate (pentasaccharide 4′‐phosphate) and α‐Kdo‐(2–8)‐α‐Kdo‐(2–4)‐α‐Kdo‐(2–6)‐β‐D‐Glc p N‐(1–6)‐α/β‐D‐Glc p N 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‐atom‐bombardment mass spectrometry.