An alternate pattern for globoside oligosaccharide expression in Haemophilus influenzae lipopolysaccharide: structural diversity in nontypeable strain 1124.
Common structural motifs of Haemophilus influenzae lipopolysaccharide (LPS) are globotetraose [beta-d-GalpNAc-(1-->3)-alpha-d-Galp-(1-->4)-beta-d-Galp-(1-->4)-beta-d-Glcp] and its truncated versions globoside [alpha-d-Galp-(1-->4)-beta-d-Galp-(1-->4)-beta-d-Glcp] and lactose [beta-d-Galp-(1-->4)-beta-d-Glcp] linked to the terminal heptose (HepIII) of the triheptosyl inner-core moiety l-alpha-d-Hepp-(1-->2)-[PEA-->6]-l-alpha-d-Hepp-(1-->3)-l-alpha-d-Hepp-(1-->5)-[PPEA-->4]-alpha-Kdo-(2-->6)-lipid A. We report here structural studies of LPS from nontypeable H. influenzae strain 1124 expressing these motifs linked to both the proximal heptose (HepI) and HepIII at the same time. This novel finding was obtained by structural studies of LPS using NMR techniques and electrospray ionization mass spectrometry (ESI-MS) on O-deacylated LPS and core oligosaccharide material (OS) as well as ESI-MS(n)() on permethylated dephosphorylated OS. The use of defined mutants allowed us to confirm structures unambiguously and understand better the biosynthesis of each of the globotetraose units. We found that lgtC is involved in the expression of alpha-d-Galp-(1-->4)-beta-d-Galp in both extensions, whereas lic2A directs only the expression of beta-d-Galp-(1-->4)-beta-d-Glcp when linked to HepIII. The LPS of NTHi strain 1124 contained sialylated glycoforms that were identified by CE-ESI-MS/MS. A common sialylated structure in H. influenzae LPS is sialyllactose linked to HepIII. This structure exists in strain 1124. However, results for the lpsA mutant indicate that sialyllactose extends from HepI as well, a molecular environment for sialyllactose in H. influenzae that has not been reported previously. In addition, the LPS was found to carry phosphorylcholine, O-linked glycine, and a third PEA group which was linked to O3 of HepIII.
Yildirim
Håkan H
HH
Clinical Research Centre, Karolinska Institutet and University College of South Stockholm, NOVUM, S-141 86 Huddinge, Sweden.
Li
Jianjun
J
Richards
James C
JC
Hood
Derek W
DW
Moxon
E Richard
ER
Schweda
Elke K H
EK
D001426:Bacterial Proteins / Q000235:genetics; D002240:Carbohydrate Sequence; D015870:Gene Expression; D005798:Genes, Bacterial; D005915:Globosides / Q000737:chemistry* / Q000235:genetics; D006193:Haemophilus influenzae / Q000737:chemistry* / Q000145:classification / Q000235:genetics; D008070:Lipopolysaccharides / Q000737:chemistry*; D008969:Molecular Sequence Data; D009154:Mutation; D019906:Nuclear Magnetic Resonance, Biomolecular; D009844:Oligosaccharides / Q000737:chemistry / Q000235:genetics; D021241:Spectrometry, Mass, Electrospray Ionization
D001426:Bacterial Proteins; D005915:Globosides; D008070:Lipopolysaccharides; D009844:Oligosaccharides; C102237:lic2A protein, Haemophilus influenzae