Lipopolysaccharides methylation analysis

Terminal 3,6-dideoxyhexoses that occur in lipopolysaccharides from Salmonella and Yersinia (Pasteurella) species7 could be hydrolyzed off with a high degree of selectivity. They may, therefore, be located by methylation analysis of the original lipopolysaccharide and of a partially hydrolyzed sample. Thus, for the Salmonella typhi-murium 395 MS lipopolysaccharide, composed18 of oligosaccharide... 

A. Gamian, E. Romanowska, A. Romanowska, C. Lugowski, J. Dabrowski, and K. Trauner, Citrobacter lipopolysaccharides Structure elucidation of the O-specific polysaccharide from strain PCM 1487 by mass spectrometry, one-dimensional and two-dimensional H NMR spectroscopy and methylation analysis, Eur. J. Biochem., 146 (1985) 641-647. 

From this methylation analysis and other studies, the complete structure for the O-specific side-chains of the lipopolysaccharide from S. typhi was postulated to be that shown in 29. 

The chemical structure of lipid A of lipopolysaccharide isolated from Comamonas testosteroni was recently determined by lida et al. (1996) by means of methylation analysis, mass spectrometry and NMR. The lipid A backbone was found to consist of 6-0-(2-deoxy-2-amino-P-D-glucopyrano-syl)-2-deoxy-2-amino-alpha-D-glucose which was phosphorylated in positions 1 and 4. Hydroxyl groups at positions 4 and 6 were unsubstituted, and position 6 of the reducing terminal residue was identified as the attachment site of the polysaccharide group. Fatty acid distribution analysis and ES/MS of lipid A showed that positions 2,2, 3 and 3 of the sugar backbone were N-acylated or O-acylated by R-3-hydroxydecanoic acid and that the hydroxyl groups of the amide-linked residues attached to positions 2 and 2 were further O-acylated by tetradecanoic and dodecanoic acids, respectively. 

Studies of the chemical composition and biology of the lipopolysaccharide from Anacystis nidulans have been reported. The glycan component of this lipopolysaccharide is composed mainly of residues of o-mannose and small proportions of 3- and 4-0-methyl-D-mannose, D-galactose, D-glucose, L-fucose, 2-amino-2-deoxy-D-glucose, 2-amino-2-deoxy-D-mannose, and 3-deoxy-2-octulo-sonic acid. Methylation analysis and oxidation of the D-mannan chain with periodate revealed the presence of (1 3)- and (1 4)-linkages in the ratio 3 1. 

The lipopolysaccharides have been isolated from two wild-type strains of E. coli K12, two core-deficient mutants, and an SR recombinant with Salmonella typhimurium Methylation analysis of the oligosaccharides released on dephosphorylation of the lipopolysaccharides revealed differences in the extents of completion of the core structures. The non-reducing end of the complete core structure of lipopolysaccharides from the E. coli K12 wild-type strains is substituted with either 2-acetamido-2-deoxy-D-glucose or, possibly, 2-acetamido-2-deoxy-D-mannopyranosyluronic acid, whereas the complete K12 core in the SR recombinant is substituted with an S-specific oligosaccharide of S. typhimurium. These substituents are attached to 0-6 of the D-glucopyranosyl residue at the non-reducing end of the core oligosaccharide, as shown in (8). 

UDP-D-galactose to the terminal, non-reducing D-glucosyl residues of these oligosaccharides. The tetrasaccharide repeating unit (8) of the O-specific side-chains of the cell-wall lipopolysaccharide from E. colt 075 has been established by methylation analysis and Smith degradation. ... 

The O-specific polysaccharide of the lipopolysaccharide from Yersinia pseudotuberculosis type IB contains residues of o-mannose, L-fucose, 3,6-dideoxy-D-r/6o-hexose (paratose), and 2-acetamido-2-deoxy-D-glucose. The repeating unit (12) of the polysaccharide was assigned on the basis of methylation analysis,... 

Note The important difference between the alkylation procedures involving sodium hydride in polar apiotic solvents and those involving alkyl halides and metal salts is that 0-acyl groups are completely replaced by 0-alkyl groups [30], N-Acyl groups survive and N-ulkyl-N-acylamido derivatives are produced. This alkylation can be performed on an extremely small amount of material and, thus, the procedure provides a rapid means for methylation linkage analysis in the polysaccharide [30], lipopolysaccharide [31] and peptide [20, 32] fields. 

The capillary g.c.-e.i.- and c. i.(NH2)-m.s. of partially methylated and acetylated derivatives of 3-deoxy-2-keto-aldonlc acids and 3-deoxyaldltols has been reported in preparation for the methylatlon analysis of KDO units in bacterial lipopolysaccharides." In the c. i. ( 1-C H q )-m. s. of some branched-chain hexopyranoside derivatives (with one or two Ac, C02Me, CONH or CN groups at C-3), the [M+C Hg]" and several primary fragment ion intensities were found to be dependent upon the nature and stereochemistry of the C-3 substituent. ... 

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