Elucidation of the structure of the lipopolysaccharide core and the linkage between the core and the O-antigen in Pseudomonas aeruginosa immunotype 5 using strong alkaline degradation of the lipopolysaccharide.

Abstract

The products of the strong alkaline degradation of the lipopolysaccharide (LPS) of Pseudomonas aeruginosa immunotype 5 were separated by anion-exchange HPLC and studied by electrospray ionization mass spectrometry and NMR spectroscopy. It was found that two major products have the same inner core region and lipid A carbohydrate backbone (A) but different outer core regions (B and C). The difference is in the position of a rhamnose residue, which is substituted with either an additional glucose residue (B) or a disaccharide remainder of the degraded O-polysaccharide (C). The site and the configuration of the linkage between the O-polysaccharide and the core were determined and, together with published data, the structure of the so-called biological repeating unit of the O-antigen was defined (D). The glycosidic linkage of the quinovosamine residue is beta when it links the O-polysaccharide to the core (C) and alpha when it connects the interior repeating units of the O-polysaccharide to each other (D) [Formula: see text]. In the structures shown Rha stands for rhamnose, Kdo for 3-deoxy-D-manno-oct-2-ulosonic acid, Hep for L-glycero-D-manno-heptose, GalNAcA for 2-acetamido-2-deoxygalacturonic acid, QuiN for 2-amino-2,6-dideoxyglucose (quinovosamine), DeltaHexNA for 2-amino-2-deoxy-D-threo-hex-4-enuronic acid; all monosaccharides are in the pyranose form and have the D configuration, except for Rha and GalNAcA that have the L configuration. In C, the remainder of the degraded O-polysaccharide is shown in bold type.