E.K.H. Schweda <Elke.Schweda
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-Hepp-(1→2)-[PEtn→6]-L-α-D-Hepp-(1→3)-L-α-D-Hepp-(1→5)-[PPEtn→4]-α-Kdo-(2→6)-Lipid A] with PCho→6)-β-D-Glcp (GlcI) substituting the proximal heptose (HepI) at O-4. Strain 1247 expresses the common structural motifs of H. influenzae; globotetraose [β-D-GalpNAc-(1→3)-α-D-Galp-(1→4)-β-D-Galp-(1→4)-β-D-Glcp-(1→] and its truncated versions globoside [α-D-Galp-(1→4)-β-D-Galp-(1→4)-β-D-Glcp-(1→] and lactose [β-D-Galp-(1→4)-β-D-Glcp-(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-Gal-(1→4)-β-D-Glcp-(1→] as an extension from GlcI which is a novel finding for NTHi LPS.
13C NMR, 1H NMR, methylation, NMR-2D, GC-MS, HF solvolysis, ESI-MS, mild acid hydrolysis, de-O-acylation with hydrazine, composition analysis, NMR-1D, alkaline deacylation, HPAEC-PAD, CE-ESI-MS/MS, CE-ESI-MS, LC-ESI-MS
HexNAcHex4 glycoform core oligosaccharide from mutant strain H. influenzae (NTHi) 1247lpsA
13C NMR data:
Linkage Residue C1 C2 C3 C4 C5 C6 C7
5,3,2,? Ac
5,3,2 aXLDmanHepp 101.7 70.9 69.4 ? ? ? ?
5,3,6,0 xXEtN
5,3,6 P
5,3 aXLDmanHepp 98.6 79.2 ? ? ? 75.3 ?
5,4,4,4,4,3,2 Ac ? 22.6
5,4,4,4,4,3 bDGalpN 103.8 54.1 70.3 66.6 ? ?
5,4,4,4,4 aDGalp 100.9 69.0 79.0 69.2 ? ?
5,4,4,4 bDGalp 103.7 70.9 72.3 77.5 75.8 ?
5,4,4 bDGlcp 103.1 73.5 74.9 78.7 75.9 59.8
5,4,6,0 xXCho ? ? 53.24
5,4,6 P
5,4,? Ac ? 23.6
5,4 bDGlcp 103.2 73.5 75.0 78.7 71.8 64.1
5 aXLDmanHepp 97.5 70.8 72.16 73.9 ? 68.0 ?
x?Sug?
1H NMR data:
Linkage Residue H1 H2 H3 H4 H5 H6 H7
5,3,2,? Ac
5,3,2 aXLDmanHepp 5.20 4.07 3.92 ? ? ? ?
5,3,6,0 xXEtN
5,3,6 P
5,3 aXLDmanHepp 5.82 4.27 ? ? 3.88 4.57 3.73
5,4,4,4,4,3,2 Ac - 2.05
5,4,4,4,4,3 bDGalpN 4.64 3.94 3.75 3.94 3.70 ?
5,4,4,4,4 aDGalp 4.93 3.91 3.97 4.26 ? ?
5,4,4,4 bDGalp 4.54 3.60 3.76 4.04 3.81 ?
5,4,4 bDGlcp 4.68 3.31 3.66 3.72 3.72 3.90
5,4,6,0 xXCho ? ? 3.24
5,4,6 P
5,4,? Ac - 1.92
5,4 bDGlcp 4.57 3.54 3.61 3.71 3.72 4.31
5 aXLDmanHepp 5.05 3.97 3.99 4.29 ? 4.11 ?
x?Sug?
1H/13C HSQC data:
Linkage Residue C1/H1 C2/H2 C3/H3 C4/H4 C5/H5 C6/H6 C7/H7
5,3,2,? Ac
5,3,2 aXLDmanHepp 101.7/5.20 70.9/4.07 69.4/3.92 ?/? ?/? ?/? ?/?
5,3,6,0 xXEtN
5,3,6 P
5,3 aXLDmanHepp 98.6/5.82 79.2/4.27 ?/? ?/? ?/3.88 75.3/4.57 ?/3.73
5,4,4,4,4,3,2 Ac 22.6/2.05
5,4,4,4,4,3 bDGalpN 103.8/4.64 54.1/3.94 70.3/3.75 66.6/3.94 ?/3.70 ?/?
5,4,4,4,4 aDGalp 100.9/4.93 69.0/3.91 79.0/3.97 69.2/4.26 ?/? ?/?
5,4,4,4 bDGalp 103.7/4.54 70.9/3.60 72.3/3.76 77.5/4.04 75.8/3.81 ?/?
5,4,4 bDGlcp 103.1/4.68 73.5/3.31 74.9/3.66 78.7/3.72 75.9/3.72 59.8/3.90
5,4,6,0 xXCho ?/? ?/? 53.24/3.24
5,4,6 P
5,4,? Ac 23.6/1.92
5,4 bDGlcp 103.2/4.57 73.5/3.54 75.0/3.61 78.7/3.71 71.8/3.72 64.1/4.31
5 aXLDmanHepp 97.5/5.05 70.8/3.97 72.16/3.99 73.9/4.29 ?/? 68.0/4.11 ?/?
x?Sug?
report error
Found 1 record.
Displayed record 1
, Life stage: adult]; chronic respiratory tract infections [ICD11: CA45 
ki.se>