Genetic and Structural Characterization of L11 Lipooligosaccharide from Neisseria meningitidis Serogroup A Strains

Abstract

The lipooligosaccharide (LOS) of immunotype L11 is unique within serogroup A meningococci. In order to resolve its molecular structure, we conducted LOS genotyping by PCR analysis of genes responsible for ␣-chain sugar addition (lgtA, -B, -C, -E, -H, and -F) and inner core substituents (lgtG, lpt-3, and lpt-6). For this study, we selected seven strains belonging to subgroup III, a major clonal complex responsible for meningococcal meningitis epidemics in Africa. In addition, we sequenced the homopolymeric tract regions of three phase-variable genes (lgtA, lgtG, and lot-3) to predict gene functionality. The fine structure of the L11 LOS of each strain was determined using composition and glycosyl linkage analyses, NMR, and mass spectrometry. The masses of the dephosphorylated oligosaccharides were consistent with an oligosaccharide composed of two hexoses, one N-acetyl-hexosamine, two heptoses, and one KDO, as proposed previously. The molar composition of LOS showed two glucose residues to be present, in agreement with lgtH sequence prediction. Despite phosphoethanolaminetransferase genes lpt-3 and lpt-6 being present in all seven Neisseria meningitidis strains, phosphoethanolamine (PEtn) was found at both O-3 and O-6 of HepII among the three ST-5 strains, whereas among the four ST-7 strains, only one PEtn was found and located at O-3 of the HepII. The L11 LOS was found to be O-acetylated, as was indicated by the presence of the lot-3 gene being in-frame in all of the seven N. meningitidis strains. To our knowledge, these studies represent the first full genetic and structural characterization of the L11 LOS of N. meningitidis. These investigations also suggest the presence of further regulatory mechanisms affecting LOS structure microheterogeneity in N. meningitidis related to PEtn decoration of the inner core.

Antigenic Potential of a Highly Conserved Neisseria meningitidis Lipopolysaccharide Inner Core Structure Defined by Chemical Synthesis

“…The immunotype expression is subjected to phase variation (Berrington et al, 2002;Zhu et al, 2002). L1-L8 immunotypes are associated with serogroups B and C, and L9 immunotype is common in serogroups A, B, and C, whereas L10-L12 immunotypes are found in serogroup A (Mistretta et al, 2010). In contrast, the LPS inner core is highly conserved because inner core glycosyltransferase genes are not subjected to phase variation (Kahler et al, 2005).…”

Importance of Antibodies to Lipopolysaccharide in Natural and Vaccine-Induced Serum Bactericidal Activity against Neisseria meningitidis Group B

“…Three immunotypes (L9, L10, and L11) were associated mostly with serogroup A strains, and the other eight were associated with all other serogroups. These immunotypes were subsequently confirmed by structural analysis of the respective oligosaccharides (9,16,20,24,29,37,38,42). Since that time, it has become evident that additional variations in structure, not identified by the initial immunotyping scheme, such as the substitution of glycine at the 7 position of Hep II, occur among N. meningitidis strains (25,44).…”

Improvement of Immunogenicity of Meningococcal Lipooligosaccharide by Coformulation with Lipidated Transferrin-Binding Protein B in Liposomes: Implications for Vaccine Development

“…The OSs were separated isocratically using 80 mM NaOAc in 75 mM NaOH for 15 min followed by a linear gradient to 400 mM NaOAc in 75 mM NaOH over 50 min. The localization of phosphoethanolamine (PEA) substituents was determined by nu-clear magnetic resonance (NMR) spectroscopy as previously described (49).…”