Abstract

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The structures of core regions of lipopolysaccharides from Escherichia coli

The structures of the core types R3 and R4 were completed ( Müller-Loennies et al. , 2002 ). The inner core structure of the latter is very similar to that of the core types R-2 and K-12; the structure of the last of which has also been completely elucidated ( Müller-Loennies et al. , 2003b ). Here, a novel inner core structure was identified, which was accompanied by a truncation of the outer core, thus indicating that inner core structural changes influence outer structures. Also, it was found that overexpression of the waaZ gene (implicated to encode the transfer of Kdo III to Kdo II in S. enterica and E. coli R2 and K-12) leads to truncation of the outer core and to reduction of the amount of O-antigen ( Frirdich et al. , 2003 ). In the investigation by Müller-Loennies (2003b) , the waaZ gene was not overexpressed and thus could not have been the reason for the identified truncation; however, the transfer of Kdo III to Kdo II could have been due to the gene product WaaZ. Finally, isolated core oligosaccharides from E. coli core types R1-R4 and S. enterica core types R1S and R2S lipopolysaccharides, and from the E. coli R3 core type mutant strain J-5 were utilized to identify the cross-reactive epitope of the cross-protective monoclonal antibody WN1 222-5, which is mainly determined by Hep III and the phosphate group at O-4 of Hep II ( Müller-Loennies et al. , 2003a ).

Core oligosaccharides different to the S. enterica type

This core type possesses as common partial structure l , d -Hep-(1→7)- l , d -Hep-(1→3)- l , d -Hep-(1→5)-Kdo, which is not substituted at O-3 of Hep II by Glc and in which heptose residues are not generally phosphorylated. Position O-4 of Hep I is substituted by a hexose residue or oligosaccharide.

Klebsiella pneumoniae ( Table 2 )

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The structures of core regions of lipopolysaccharides from Klebsiella pneumoniae

Presently, three core types of lipopolysaccharides of K. pneumoniae have been identified ( Vinogradov & Perry, 2001 ; Holst, 2002 ; Regué, 2005b ), the latest of which was identified in strain 52145 (serotypes O1:K2). This core region does not possess the disaccharide l , d -Hep-(1→4)-α-Kdo that substitutes O-6 of the outer core Glc N residue and which was identified in most O-serotypes investigated so far. Instead, the Glc N is substituted at O-4 by a β-Glc-(1→6)-Glc disaccharide. Also, the β-Glc residue substituting Hep I at O-4 is not further substituted ( Regué, 2005a , b ). For a number of O-serotypes possessing the disaccharide l , d -Hep-(1→4)-Kdo that substitutes O-6 of the outer core Glc N , the position of the O-antigen linkage was determined, which was in all cases O-5 of this particular Kdo residue ( Vinogradov et al. , 2002c ). With regard to biosynthesis, the gene products WabH, WabI and WabJ were identified, responsible for the synthesis of the outer core l , d -Hep-(1→4)-Kdo-(2→6)-Glc N sequence ( Frirdich et al. , 2004 ). However, since WabH was found to transfer Glc N Ac to the GalA residue of the outer core, but Glc N had been identified as outer constituent rather than Glc N Ac, another enzyme responsible for de- N -acetylation was suggested and then identified as WabN ( Regué, 2005a ). The negative charge(s) of the GalA residue(s) were found to be highly important in outer membrane integrity, equivalent to phosphate residues in other core types ( Frirdich et al. , 2005 ).

Proteus penneri ( Table 3 )

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The structures of core regions of lipopolysaccharides from Proteus penneri

In continuation of earlier highly significant work ( Vinogradov et al. , 2002a ), the core regions of lipopolysaccharides from various serotypes of Proteus penneri have been structurally characterized ( Vinogradov & Sidorczyk, 2002 ; Vinogradov et al. , 2002c ). All core structures possess as common carbohydrate backbone l , d -Hep-(1→7)- l , d -Hep-(1→3)- l , d -Hep-(1→5)-[Kdo-(2→4)-]Kdo. Hep I is substituted at O-4 by a terminal β-Glc residue, and Hep II is substituted by GalA at O-3 and 2-aminoethyl phosphate at O-6. This backbone is differently substituted determining the structural specificities of these cores.

Yersinia ( Table 4 )

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The structures of core regions of lipopolysaccharides from Yersinia

In the past 5 years, the general structure of the core region of lipopolysaccharides from Y. pestis ( Anisimov et al. , 2004 ) was elucidated and variations due to temperature changes and interspecies diversity were characterized ( Vinogradov et al. , 2002 ; Knirel et al. , 2005a , b ). The structure is similar to that of the lipopolysaccharides core region of Yersinia enterocolitica O:8 ( Oertelt et al. , 2001 ) possessing the backbone l , d -Hep-(1→7)- l , d -Hep-(1→3)- l , d -Hep-(1→5)-[Kdo-(2→4)-]Kdo, which is substituted at O-3 of Hep II by β-Glc N Ac and at O-7 of Hep III either by β-Gal or by d , d -Hep, all in nonstoichiometric amounts. These substituents may vary with temperature change and in different subspecies. Furthermore, the amino acid glycine was identified, but could not be localized. Most strikingly was the finding that Kdo II may partially be replaced by Ko. Such Ko-(2→4)-Kdo structural moiety had earlier been identified only in lipopolysaccharides of the genus Burkholderia and was thought to be specific here. Although this core region is generally not phosphorylated, Ko or Kdo II are substituted at O-7 by 2-aminoethanol phosphate in lipopolysaccharides of bacteria grown at 6°C. Such substitution had been identified earlier in lipopolysaccharides from S. enterica and E. coli (grown at 37°C) ( Holst et al. , 1990 ).

The core region of Y. enterocolitica O:3 has been revised (own unpublished work). Not d -Fuc p NAc but 2-acetamido-2,6-dideoxy- d - xylo -hexos-4-ulopyranose was found to link the outer core region to the inner core.

Serratia marcescens ( Table 5 )

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The structures of core regions of lipopolysaccharides from Serratia marcescens

Serratia marcescens is a nosocomial human pathogen (pneumonia, meningitis, urinary tract diseases). Two lipopolysaccharides core structures have been published so far ( Coderch et al. , 2004 ; Vinogradov et al. , 2006 ), which both combine features of the core regions of lipopolysaccharides from Y. pestis , Proteus mirabilis/Proteus penneri , K. pneumoniae and Burkholderia cepacia/Burkholderia pseudomallei . The cores of strains 111R (serotype O:29) and IFO 3735 were highly heterogenous.

Plesiomonas shigelloides ( Table 6 )

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The structures of core regions of lipopolysaccharides from Plesiomonas shigelloides

Plesiomonas shigelloides is a rod-shaped bacterium, which has been isolated from freshwater, freshwater fish, shellfish and from a great number of animals. It causes diarrhea in humans. The core structures of lipopolysaccharides from Plesiomonas shigelloides serotypes O:54, O74 and O:13 have been published ( Kasbowska et al. , 2006 ; Lukasiewicz et al. , 2006a , b ; Niedziela et al. , 2006 ). All three core regions were free of phosphate residues. The core structures of serotypes O:13 and O:54 possessed, as novel feature, a Gal residue that is β-(1→4)-linked to Hep I. In the core region of serotype O:74, a GalA-(1→6)-β-Glc disaccharide substitutes this position. The linkage points of the O-specific polysaccharides were also determined. Whereas in serotype O:54 lipopolysaccharides the O-antigen is linked to a branched oligosaccharide that substitutes O-3 of Hep II, the O:13 antigen is linked via a GalA residue to O-7 of Hep III. The O:74 antigen is also linked via a GalA residue, however, to O-3 of Hep II. Here, the GalA is further substituted with d , d -Hep at O-2.

Concluding remarks

In the past 5 years, there has again been considerable progress in the structural elucidation of core regions of enterobacterial lipopolysaccharides obtained from various bacterial species. Core regions comprise as partial structure the tetrasaccharide l , d -Hep-(1→7)- l , d -Hep-(1→3)- l , d -Hep-(1→5)-Kdo, and if core structures from one bacterial family are compared, a common structural theme is often identified, which may be varied in different species by different substituents. Apart from the finding that a mutant of E. coli K-12 has been identified that is viable possessing only lipid IV _A as a minimal ‘lipopolysaccharides’ structure ( Meredith et al. , 2006 ), the common principle of lipopolysaccharides may still be seen as the core oligosaccharide and the lipid A, as identified in all other lipopolysaccharides investigated so far. The expression of a (O-specific) polysaccharide in lipopolysaccharides is not a prerequisite for bacterial survival; however, the finding that the polysaccharide portion in S-form lipopolysaccharides may be furnished either by the O-chain or a capsule or ECA indicates that such lipopolysaccharides form is highly advantageous in many bacteria. In enterobacterial lipopolysaccharides, the binding of the core region to lipid A occurs always via a Kdo residue, and, as in all other lipopolysaccharides, the core region is always negatively charged (provided by phosphoryl substituents and/or sugar acids like Kdo and uronic acids) which is thought to contribute to the rigidity of the Gram-negative cell wall through intermolecular cationic cross-links ( Frirdich et al. , 2005 ).

In enterobacterial lipopolysaccharides, all core structures contain heptoses, either l , d -Hep alone or d , d -Hep in addition to it. If present, d , d -Hep either decorates the inner core region (e.g. in Y. enterocolitica ) or is attached to more remote parts of the carbohydrate chain (in one core type of K. pneumoniae lipopolysaccharides). Still, the regulation of the distribution of l , d -Hep and d , d -Hep in the core region is not understood, and it is not known whether l , d -Hep and d , d -Hep are transferred by different transferases.

Acknowledgements

This research is supported by the Deutsche Forschungsgemeinschaft (grants SFB-TR 22 A1 and A2) and the European Union (GABRIEL, GALTRAIN, GA ² LEN).

References