Expression of a new disialyllacto structure in the lipopolysaccharide of non-typeable Haemophilus influenzae.

Abstract The structure of lipopolysaccharide (LPS) expressed by non-typeable Haemophilus influenzae (NTHi) strains 1008 and 1247 has been investigated by mass spectrometry and NMR analyses on O-deacylated LPS and core oligosaccharide material. Both strains express the conserved triheptosyl inner core, [ l -α- d -Hep p -(1→2)-[ P Etn→6]- l -α- d -Hep p -(1→3)- l -α- d -Hep p -(1→5)-[ PP Etn→4]-α-Kdo-(2→6)-Lipid A] with P Cho→6)-β- d -Glc p (GlcI) substituting the proximal heptose (HepI) at O-4 . Strain 1247 expresses the common structural motifs of H. influenzae ; globotetraose [β- d -Gal p NAc-(1→3)-α- d -Gal p -(1→4)-β- d -Gal p -(1→4)-β- d -Glc p -(1→] and its truncated versions globoside [α- d -Gal p -(1→4)-β- d -Gal p -(1→4)-β- d -Glc p -(1→] and lactose [β- d -Gal p -(1→4)-β- d -Glc p -(1→] linked to the terminal heptose of the inner core and GlcI. A genetically distinct NTHi strain, 1008, expresses identical structures to strain 1247 with the exception that it lacks GalNAc. A lpsA mutant of strain 1247 expressed LPS of reduced complexity that facilitated unambiguous structural determination of the oligosaccharide from HepI. By CE-ESI-MS/MS we identified disialylated glycoforms indicating disialyllactose [α-Neu5Ac-(2→8)-α-Neu5Ac-(2→3)-β- d -Ga l -(1→4)-β- d -Glc p -(1→] as an extension from GlcI which is a novel finding for NTHi LPS.