Structure of the exceptionally large nonrepetitive carbohydrate backbone of the lipopolysaccharide of Pectinatus frisingensis strain VTT E‐82164

Abstract

The structures of the oligosaccharides obtained after acetic acid hydrolysis and alkaline deacylation of the rough-type lipopolysaccharide (LPS) from Pectinatus frisingensis strain VTT E-82164 were analysed using NMR spectroscopy, MS and chemical methods. The LPS contains two major structural variants, differing by a decasaccharide fragment, and some minor variants lacking the terminal glucose residue. The largest structure of the carbohydrate backbone of the LPS that could be deduced from experimental results consists of 25 monosaccharides (including the previously found Ara4NP residue in lipid A) arranged in a well-defined nonrepetitive structure:We presume that the shorter variant with R 1 ¼ H represents the core-lipid A part of the LPS, and the additional fragment is present instead of the O-specific polysaccharide. Structures of this type have not been previously described. Analysis of the deacylation products obtained from the LPS of the smooth strain, VTT E-79100 T , showed that it contains a very similar core but with one different glycosidic linkage.

The structure of the carbohydrate backbone of the lipopolysaccharide of Pectinatus frisingensis strain VTT E-79104

“…The oligosaccharide 1 had most of its structure identical to that of the respective oligosaccharide from strain E-82164, and differed by the absence of the K-J (Scheme 1) fragment and by the substitution of the Man G with a-Gal at position 2 instead of the substitution with a-Glc at position 3. 5 The 1 H NMR spectrum of oligosaccharide 2 had strong overlap of the anomeric signals and the monosaccharide residues situated close to the nonreducing end gave two series of signals. Although it had the expected mass of 3418.2 Da (dHex 1 Hex 7 Hep 2 HexN 6 -Kdo 3 P 1 Me 2 ) and one could find duplicated spin systems of all expected monosaccharides, the spectra were not completely interpreted and the reason for duplication of the signals was not found.…”

“…This indicates the presence of two structurally distinct LPS populations, as was recently shown 4 in P. frisingensis VTT E-82164. After structure determination of one such LPS (VTT E-82164), 5 we used the same approaches to the structural analysis of the strain VTT E-79104.…”

“…frisingensis VTT E-79104 was obtained from the culture collection of VTT Biotechnology, Espoo, Finland. All experiments were performed as described, 5 except that NMR spectra were recorded on a Varian UNITY INOVA 600 spectrometer, and more harsh conditions were used for alkaline deacylation (120°C, 24 h) because of high stability of the N-acetyl groups.…”

“…The inner core region was devoid of heptose and contained an additional Kdo residue outside the region of linkage to lipid A. 5 Here we present the results of structural analysis of the LPS from another strain of the O-chain-lacking type, P. frisingensis VTT E-79104, which shares a general architecture of LPS with P. frisingensis VTT E-82164.…”

Structural Analysis of a Novel Polysaccharide of the Lipopolysaccharide‐Deficient Extremophile Gram‐Negative Bacterium Thermus thermophilus HB8

“…In several examples the substituent masking the repetitiveness of the polymer is a noncarbohydrate group (i.e., an acetyl group), [19] but, recently, carbohydrate residues have been found to be unsystematically attached to a carbohydrate chain, and, in two cases, a saccharide chain with a completely unsystematic collection of carbohydrate residues was reported. [14,20] Even in the polysaccharide derived from the outer membrane of the Gram-negative bacterium Thermus thermophilus HB8, there is a repeating unit masked by a nonstoichiometric carbohydrate substituent. Its peculiarity resides in the fact that the ''substituent'' linked to the always present oligosaccharide unit is a trisaccharide chain.…”