Structure and serological characterization of the O-antigen of Proteus mirabilis O18 with a phosphocholine-containing oligosaccharide phosphate repeating unit.
A phosphorylated, choline-containing polysaccharide was obtained by O-deacylation of the lipopolysaccharide (LPS) of Proteus mirabilis O18 by treatment with aqueous 12% ammonia, whereas hydrolysis with dilute acetic acid resulted in depolymerisation of the polysaccharide chain by the glycosyl phosphate linkage. Treatment of the O-deacylated LPS with aqueous 48% hydrofluoric acid cleaved the glycosyl phosphate group but, unexpectedly, did not affect the choline phosphate group. The polysaccharide and the derived oligosaccharides were studied by NMR spectroscopy, including 2D 1H,1H COSY, TOCSY, ROESY, 1H,13C HMQC and HMQC-TOSCY experiments, along with chemical methods, and the following structure of the pentasaccharide phosphate repeating unit was established: [carbohydrate structure in text] Where ChoP=Phosphocoline Immunochemical studies of the LPS, O-deacylated LPS and partially dephosphorylated pentasaccharide using rabbit polyclonal anti-P. mirabilis O18 serum showed the importance of the glycosyl phosphate group in manifesting the serological specificity of the O18-antigen.
Fudala
Rafal
R
Institute of Microbiology and Immunology, University of Lodz, 90-237 Lodz, Poland.
Kondakova
Anna N
AN
Bednarska
Katarzyna
K
Senchenkova
Sof'ya N
SN
Shashkov
Alexander S
AS
Knirel
Yuriy A
YA
Zähringer
Ulrich
U
Kaca
Wieslaw
W
D019342:Acetic Acid / Q000737:chemistry; D000117:Acetylglucosamine / Q000032:analysis; D000641:Ammonia / Q000737:chemistry; D000818:Animals; D015153:Blotting, Western; D001894:Borohydrides / Q000737:chemistry; D002240:Carbohydrate Sequence; D002849:Chromatography, Gas; D003429:Cross Reactions / Q000276:immunology; D003641:Deamination; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000276:immunology; D005690:Galactose / Q000032:analysis; D005947:Glucose / Q000032:analysis; D006858:Hydrofluoric Acid / Q000737:chemistry; D006868:Hydrolysis; D007106:Immune Sera / Q000276:immunology; D008070:Lipopolysaccharides / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D019081:O Antigens / Q000737:chemistry* / Q000276:immunology; D025942:Oligosaccharides, Branched-Chain / Q000737:chemistry* / Q000276:immunology; D010710:Phosphates / Q000032:analysis* / Q000276:immunology; D010767:Phosphorylcholine / Q000032:analysis* / Q000276:immunology; D011513:Proteus mirabilis / Q000737:chemistry* / Q000145:classification; D011817:Rabbits; D012703:Serotyping; D021241:Spectrometry, Mass, Electrospray Ionization
D001894:Borohydrides; D000939:Epitopes; D007106:Immune Sera; D008070:Lipopolysaccharides; D019081:O Antigens; D025942:Oligosaccharides, Branched-Chain; D010710:Phosphates; D010767:Phosphorylcholine; D000641:Ammonia; D005947:Glucose; D019342:Acetic Acid; D006858:Hydrofluoric Acid; D000117:Acetylglucosamine; D005690:Galactose