Structure of the core oligosaccharide in the serotype O8 lipopolysaccharide from Klebsiella pneumoniae

Abstract

Two classes of mutants with O-antigen-deficient lipopolysaccharides were isolated from the serotype O8 reference strain, belonging to Klebsiella pneumoniae subspecies ozaenae. These mutants were selected by resistance to bacteriophage KO1-2, which recognizes and lyses strains with lipopolysaccharide molecules containing the D-galactan II O antigen. Strain RFK-11 contains a defect in O-antigen synthesis and has a complete core, including the attachment site for O antigen. This mutation is complemented by a plasmid carrying the rfb (O-antigen biosynthesis) gene cluster from the related K. pneumoniae serotype O1. In sodium dodecyl sulfatepolyacrylamide gel electrophoresis, the lipopolysaccharide from strain RFK-9 has a mobility typical of deeprough lipopolysaccharide. RFK-9 lipopolysaccharide lacks the attachment site for O antigen. Lipopolysaccharides from strains RFK-9 and RFK-11 were isolated, and their structures were determined by methylation analyses, nuclear magnetic resonance spectroscopy, and mass spectroscopy. The deduced O8 core oligosaccharide includes the partial core structure reported for the K. pneumoniae subspecies pneumoniae serotype O1 lipopolysaccharide (M. Süsskind, S. Müller-Leonnies, W. Nimmich, H. Brade, and O. Holst, Carbohydr. Res. 269:C1-C7, 1995), consistent with the possibility of a conserved core structure within the species. The core oligosaccharide differs from those of the genera Salmonella and Escherichia by the absence of a hexose-containing outer core, the lack of phosphate residues in the inner core, and the presence of galacturonic acid residues.

Genetic Characterization of theKlebsiella pneumoniae waaGene Cluster, Involved in Core Lipopolysaccharide Biosynthesis

“…Comparison of the core LPS structure in K. pneumoniae (43,45) and E. coli (20) reveals differences in both the inner and outer core structures. In bacteriocin 28b-sensitive E. coli cells, the core LPS is involved in bacteriocin binding (11).…”

“…Plasmid pBG3 and primer pairs A1 (5Ј-CGCGGATCCCACCGCAAGCTGCTGGAAAA-3Ј) and B1 (5Ј-CCCATCCACTAAACTTAAACAGCTTTTGCGGCTGCTCATT C-3Ј) and C1 (5Ј-TGTTTAAGTTTAGTGGATGGGGTGGTCAACGCGCAA TATAC-3Ј) and D1 (5Ј-CGCGGATCCTCCTTCACCAGTGATGAGGA-3Ј) were used to obtain plasmid pKO3⌬waaE containing an internally deleted waaE gene (the first 6 codons, a 7-codon tag, and the last 24 codons) flanked by 541 bp upstream and 409 bp downstream. Plasmid pNUC41 and primer pairs A2 (5Ј-CGCAGATCTCACCTGATACCCGTATTCCAC-3Ј) and B2 (5Ј-CCCATC CACTAAACTTAAACACAGCTTAATGACCAGGATCCG-3Ј) and C2 (5Ј-T GTTTAAGTTTAGTGGATGGGGCTATCAACACCAACACCGAC-3Ј) and D2 (5Ј-CGCAGATCTCGCTGGTTATCAATGGCGTTG-3Ј) (the BglII site is double-underlined in primers A2 and D2) were used to obtain plasmid pK03⌬waaQ containing an internally deleted waaQ gene (the first 21 (43,45). In K. pneumoniae RFK-11 (O8 Ϫ :K Ϫ ), only two Kdop residues were detected (42).…”

“…Comparison of the core LPS structure in K. pneumoniae (43,45) and E. coli and S. enterica (20) reveals differences in both the inner and outer core structures (Fig. 1).…”

O antigen seroepidemiology of Klebsiella clinical isolates and implications for immunoprophylaxis of Klebsiella infections

“…The only structural data for the Klebsiella core oligosaccharide are a partial structure from an O1 isolate (44) and a complete structure from a related O8 isolate (40). The larger O8 structure contains the region first identified in O1.…”

Expression Cloning of Three Rhizobium leguminosarum Lipopolysaccharide Core Galacturonosyltransferases

“…1). Mesorhizobium huakuii, another nitrogen-fixing plant endosymbiont, replaces its lipid A1-phosphate group with a GalA residue, whereas A. pyrophilus, a hyperthermophile, replaces both the lipid A1 and 4Ј-phosphates with GalA (11,17,(25)(26)(27)53). The functions of the GalA units present in the LPS of these bacteria are unknown.…”