Structural diversity in lipopolysaccharide expression in nontypeable Haemophilus influenzae. Identification of L-glycerol-D-manno-heptose in the outer-core region in three clinical isolates.
Structural elucidation of the lipopolysaccharide (LPS) from three nontypeable Haemophilus influenzae clinical isolates, 1209, 1207 and 1233 was achieved using NMR spectroscopy and ESI-MS on O-deacylated LPS and core oligosaccharide (OS) material as well as ESI-MS(n) on permethylated dephosphorylated OS. It was found that the organisms expressed a tremendous heterogeneous glycoform mixture resulting from the variable length of the OS chains attached to the common structural element of H. influenzae, L-alpha-D-Hepp-(1-->2)-[PEtn-->6]-L-alpha-D-Hepp-(1-->3)-[beta-D-Glcp-(1-->4)]-L-alpha-D-Hepp-(1-->5)-[PPEtn-->4]-alpha-Kdop-(2-->6)-Lipid A. Notably, the O-6 position of the beta-D-Glcp residue could either be occupied by PCho or L-glycero-D-manno-heptose (L,D-Hep), which is a location for L,D-Hep that has not been seen previously in H. influenzae LPS. The outer-core L,D-Hep residue was further chain elongated at the O-6 position by the structural element beta-D-GalpNAc-(1-->3)-alpha-D-Galp-(1-->4)-beta-D-Galp, or sequentially truncated versions thereof. The distal heptose residue in the inner-core was found to be chain elongated at O-2 by the globotetraose unit, beta-D-GalpNAc-(1-->3)-alpha-D-Galp-(1-->4)-beta-D-Galp-(1-->4)-beta-D-Glcp, or sequentially truncated versions thereof. Investigation of LPS from an lpsA mutant of isolate 1233 and a lic1 mutant of isolate 1209 was also performed, which aside from confirming the functions of the gene products, simplified elucidation of the OS extending from the proximal heptose (the lpsA mutant), and showed that the organism exclusively expresses LPS glycoforms comprising the outer-core l,d-Hep residue when PCho is not expressed (the lic1 mutant).
Månsson
Martin
M
Clinical Research Centre, Karolinska Institutet and University College of South Stockholm, Huddinge, Sweden.
Hood
Derek W
DW
Moxon
E Richard
ER
Schweda
Elke K H
EK
D000107:Acetylation; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D006031:Glycosylation; D006193:Haemophilus influenzae / Q000737:chemistry* / Q000302:isolation & purification; D006539:Heptoses / Q000032:analysis*; D006801:Humans; D008070:Lipopolysaccharides / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy; D008745:Methylation; D008956:Models, Chemical; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010766:Phosphorylation; D021241:Spectrometry, Mass, Electrospray Ionization
D006539:Heptoses; D008070:Lipopolysaccharides; C026312:glycero-manno-heptose