Alteration of O-specific polysaccharide structure of symbiotically defective Mesorhizobium loti mutant 2213.1 derived from strain NZP2213.

Abstract

Mesorhizobium loti mutant 2213.1 derived from the wild-type strain NZP2213 by Tn5 mutagenesis showed impaired effectiveness of symbiosis with the host plant Lotus corniculatus (Turska-Szewczuk et al., 2007 Microbiol Res, in press). The inability of lipopolysaccharide (LPS) isolated from the mutant 2213.1 strain or de-O-acetylated LPS of the parental cells to inactivate phage A1 particles implicated alterations in the LPS structure. The O-specific polysaccharide of the mutant was studied by chemical analyses along with (1)H and (13)C NMR spectroscopy, which clearly confirmed alterations in the O-chain structure. 2D NMR data showed that the mutant O-polysaccharide consists of a tetrasaccharide repeating unit containing non-substituted as well as O-acetylated or O-methylated 6-deoxytalopyranose residues. Additionally, an immunogold assay revealed a reduced number of gold particles on the mutant bacteroid cell surface, which could result from both a diminished amount of an O-antigenic determinant in mutant LPS and modifications of structural epitopes caused by alterations in O-acetylation or O-methylation of sugar residues. Western immunoblot assay of alkaline de-O-acetylated lipophilic M. loti NZP2213 LPS showed no reactivity with homologous serum indicating a role of O-acetyl groups in its O-specificity.

A Mutation in the Mesorhizobium loti oatB Gene Alters the Physicochemical Properties of the Bacterial Cell Wall and Reduces Survival inside Acanthamoeba castellanii

“…In the previous report, it was shown that the NZP2213.1 mutant is resistant to lysis of phage A1 due to the changed LPS structure, and fully O -acetylated wild-type O-PS was indicated to be a receptor for the phage [ 19 ]. In the present study, the complemented NZP2213.1(pBGoatB) strain subjected to the action of phage A1 was lysed similarly to the WT strain.…”

“…The molar ratio of the main sugars building the O-chain i.e., 6-deoxytalose (6dTal p ) and 2- O -methyl-6-deoxytalose (2-OMe-6dTal p ), in relation to N -acetylquinovosamine (QuipNAc), which is a sugar of the outer core tetrasacharide [ 19 , 21 ], was ~33:1 and 15:2, in lipophilic S-LPS of the WT and mutant NZP2213.1 strains, respectively. In turn, the molar ratio of 6dTal p and 2-OMe-6dTal p estimated at ~27:1.9 in the O-PS of the complemented NZP2213.1(pBGaotB) strain indicated the restoration of the WT proportion of sugars ( Table 1 ).…”

“…A proper amount of smooth LPS in the outer membrane of this strain had been shown earlier to be crucial for symbiosis with the host plant [ 7 ]. In the present study, to establish the determinants of M. loti – A. castellanii interactions, we used the same NZP2213.1 mutant, whose O-PS was reduced by half and was not entirely substituted with O -acetyl groups [ 19 ]. The region of this mutant interrupted by a transposon had been characterized previously and shown to display a low similarity to a membrane glycosyltransferase gene of Caulobacter crescentus (CB15, 43%/53% identity/similarity) [ 7 ].…”

“…As previously determined, the amount of O-PS in the mutant is reduced by half in comparison to the WT strain. It is composed of non-substituted as well as O -acetylated or O -methylated 6dTal p residues [ 7 , 19 ]. In turn, the O-chain of the WT strain isolated from the phenol-soluble LPS species was reported to be highly hydrophobic due to the occurrence of 2- O -acetyl-6dTal p exclusively [ 20 ].…”

Struktura lipopolisacharydów rizobiowych i ich znaczenie w procesie symbiozy

“…W preparatach polisacharydowych tych bakterii zidentyfikowano także heksozy (Man, Gal), O-metyloheksozy, heptozę i jej metylową pochodną, aminocukry, kwasy uronowe oraz podstawniki niecukrowe np. grupy acetylowe [59,60,46,[61][62][63][64][65].…”