Novel O-polysaccharide expression, as a lipid A-core-free form, in a lipopolysaccharide-core-defective mutant of Pseudomonas aeruginosa.
Pseudomonas aeruginosa PML14e is a mutant strain, isolated from strain PML14 (Homma serotype I), that is resistant to all types of R-pyocins. PML14e completely lacked glucose and rhamnose as components of the lipopolysaccharide (LPS) outer core region. Whereas the O-polysaccharide attachment site on the LPS core was considered to be absent, PML14e was agglutinable with anti-serotype-I antibodies. The O-polysaccharide of PML14e was recovered in the supernatant after ultracentrifugation of the aqueous layer from a hot phenol/water extraction. Chromatographic behaviour and chemical analysis indicated that the PML14e O-polysaccharide was not linked to the lipid A. 1H-NMR spectroscopy indicated that the structure of the PML14e O-polysaccharide was the same as that of the O-polysaccharide from PML14. The above evidence indicated that the O-polysaccharide is expressed on the cell surface of the mutant strain PML14e as the lipid A-free form. To examine the nature of the cell surface, the accessibility of monoclonal antibodies (mAbs) against cell surface antigens was tested by enzyme-linked immunosorbent assay. An anti-lipid A mAb and an anti-outer-membrane protein mAb, the epitopes for which are considered to be exposed on rough strains, bound to a greater extent to the PML14e cells than to two other LPS-core-defective rough mutants, PML14b and PML14d. Whereas these mutants appeared to have lesser defects in the LPS core, they expressed less O-polysaccharide than PML14e. The results indicated that the epitopes exposed on rough strains, such as lipid A and outer-membrane proteins, were mainly hindered by covalently linked core oligosaccharide rather than by the O-polysaccharide chain.
Yokota
Shin-Ichi
SI
Sumitomo Pharmaceuticals Research Center, Konohanaku, Osaka 554, Japan.
D000911:Antibodies, Monoclonal; D002240:Carbohydrate Sequence; D015870:Gene Expression; D008050:Lipid A / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009154:Mutation; D019081:O Antigens / Q000737:chemistry*; D010641:Phenotype; D011135:Polysaccharides, Bacterial / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D011550:Pseudomonas aeruginosa / Q000737:chemistry / Q000235:genetics* / Q000276:immunology
D000911:Antibodies, Monoclonal; D008050:Lipid A; D019081:O Antigens; D011135:Polysaccharides, Bacterial