The Structure of O-antigen from Lipopolysaccharide of Rhizobium leguminosarum 128C53 and Its Nod-Fix-Mutant

The LPS of R. leguminosarum 128C53 smr rifr (a streptomycin and rifampicin resistant strain of wild type 128C53) and its mutant ANU54 (nod-, fix-) were isolated from the bacterial pellet by hot phenol/water extraction followed by gel filtration chromatography. The O-antigen was isolated by mild-acid hydrolysis of the LPS and purified by gel filtration chromatography using a Sephadex G-50 or G-25. The following results were the same for both the parent and mutant. The composition and linkage of the O-antigen were determined by gas chromatography (GC), GC-mass spectrometry, 1H and 13C nuclear magnetic resonance spectroscopy (NMR) techniques. The data indicate that the O-antigen of the LPS from parent R. leguminosarum 128C53 and its mutant ANU54 are identical. The O-antigen contains a tetrasaccharide repeating unit. The backbone consists of one 1,3-linked-rhamnose and two 1,3-linked-fucose residues. A terminal mannose is linked to the 2-position of one of the two fucose residues. The 1H-NMR analysis indicates that all the glycosyl residues are alpha linked. The exact position of the mannose residue is under further investigation