polygalacturonic-acid and Klebsiella-Infections

The core lipopolysaccharide (LPS) of Klebsiella pneumoniae contains two galacturonic acid (GalA) residues, but only one GalA transferase (WabG) has been identified. Data from chemical and structural analysis of LPS isolated from a wabO mutant show the absence of the inner core beta-GalA residue linked to L-glycero-D-manno-heptose III (L,D-Hep III). An in vitro assay demonstrates that the purified WabO is able to catalyze the transfer of GalA from UDP-GalA to the acceptor LPS isolated from the wabO mutant, but not to LPS isolated from waaQ mutant (deficient in l,d-Hep III). The absence of this inner core beta-GalA residue results in a decrease in virulence in a capsule-dependent experimental mouse pneumonia model. In addition, this mutation leads to a strong reduction in cell-bound capsule. Interestingly, a K66 Klebsiella strain (natural isolate) without a functional wabO gene shows reduced levels of cell-bound capsule in comparison to those of other K66 strains. Thus, the WabO enzyme plays an important role in core LPS biosynthesis and determines the level of cell-bound capsule in Klebsiella pneumoniae.

Topics: Animals; Bacterial Capsules; Carbohydrate Sequence; Electrophoresis, Polyacrylamide Gel; Female; Gas Chromatography-Mass Spectrometry; Genes, Bacterial; Glycosyltransferases; Hexuronic Acids; Klebsiella Infections; Klebsiella pneumoniae; Lipopolysaccharides; Magnetic Resonance Spectroscopy; Mice; Microscopy, Immunoelectron; Molecular Sequence Data; Mutation; Phenotype; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Virulence

The complete structures of LPS core types 1 and 2 from Klebsiella pneumoniae have been described by other authors. They are characterized by a lack of phosphoryl residues, but they contain galacturonic acid (GalA) residues, which contribute to the necessary negative charges. The presence of a capsule was determined in core-LPS non-polar mutants from strains 52145 (O1 : K2), DL1 (O1 : K1) and C3 (O8 : K66). O-antigen ligase (waaL) mutants produced a capsule. Core mutants containing the GalA residues were capsulated, while those lacking the residues were non capsulated. Since the proteins involved in the transfer of GalA (WabG) and glucosamine residues (WabH) are known, the chemical basis of the capsular-K2-cell-surface association was studied. Phenol/water extracts from K. pneumoniae 52145DeltawabH waaL and 52145DeltawaaL mutants, but not those from from K. pneumoniae 52145DeltawabG waaL mutant, contained both LPS and capsular polysaccharide, even after hydrophobic chromatography. The two polysaccharides were dissociated by gel-filtration chromatography, eluting with detergent and metal-ion chelators. From these results, it is concluded that the K2 capsular polysaccharide is associated by an ionic interaction to the LPS through the negative charge provided by the carboxyl groups of the GalA residues.

Topics: Animals; Bacterial Capsules; Carbohydrate Sequence; Hexuronic Acids; Humans; Klebsiella Infections; Klebsiella pneumoniae; Lipopolysaccharides; Mice; Mice, Inbred ICR; Molecular Sequence Data; Pneumonia, Bacterial