The structures of oligosaccharide bisphosphates isolated from the lipopolysaccharide of a recombinant Escherichia coli strain expressing the gene gseA [3-deoxy-D-manno-octulopyranosonic acid (Kdo) transferase] of Chlamydia psittaci 6BC.
The lipopolysaccharide from the recombinant strain Escherichia coli F515-140 containing the cloned gene gseA [3-deoxy-D-manno-octulopyranosonic acid (Kdo) transferase] from Chlamydia psittaci 6BC was isolated and sequentially de-O-acylated and de-N-acylated. The products were separated by high-performance anion-exchange chromatography into three fractions, two of which contained a single compound. Their structures were elucidated by high-field NMR spectroscopy as alpha-Kdo-(2-->4)-alpha-Kdo-(2-->6)-beta-D-GlcN-(1-->6)-alpha-D-GlcN 1,4'-P2 (compound 1) (tetrasaccharide bisphosphate) [Holst, O., Broer, W., Thomas-Oates, J. E., Mamat, U. & Brade, H. (1993) Eur. J. Biochem. 214, 703-710] and alpha-Kdo-(2-->4)-[alpha-Kdo-(2-->8)-]-alpha-Kdo-(2-->4)-alpha-Kdo- (2-->6)-beta-D-GlcN-(1-->6)-alpha-D-GlcN 1,4'-P2 (compound 4) (hexasaccharide bisphosphate). The third fraction comprised two pentasaccharide bisphosphates, which could be separated by affinity chromatography using an immobilized monoclonal antibody specific for the trisaccharide alpha-Kdo-(2-->8)-alpha-Kdo-(2-->4)-alpha-Kdo. The bound fraction was identified as alpha-Kdo-(2-->8)-alpha-Kdo-(2-->4)-alpha-Kdo-(2-->6)-beta-D- GlcN-(1-->6)-alpha-D-GlcN 1,4'-P2 (compound 2) [Holst, O., Broer, W., Thomas-Oates, J. E., Mamat, U. & Brade, H. (1993) Eur. J. Biochem. 214, 703-710], whereas the unbound fraction was identified as alpha-Kdo-(2-->4)-alpha-Kdo-(2-->4)-alpha-Kdo-(2-->6)-beta-D-GlcN-(1-->6 )- alpha-D-GlcN 1,4'-P2 (compound 3). This novel Kdo tetrasaccharide extends our knowledge on multifunctional Kdo transferases.
Holst
O
O
Division of Biochemical Microbiology, Forschungsinstitut Borstel, Germany.
Bock
K
K
Brade
L
L
Brade
H
H
D002240:Carbohydrate Sequence; D002689:Chlamydia / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D008070:Lipopolysaccharides / Q000737:chemistry*; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009844:Oligosaccharides / Q000737:chemistry* / Q000302:isolation & purification; D014166:Transferases / Q000096:biosynthesis / Q000235:genetics*
D008070:Lipopolysaccharides; D009844:Oligosaccharides; D014166:Transferases; C019917:3-deoxy-D-manno-octulosonate transferase