Vaccine and Emerging Infections Research, Human Health Therapeutics Research Centre, National Research Council, Ottawa, ON, K1A 0R6, Canada
Fusobacterium nucleatum is a gram-negative bacterium, part of the normal human microflora. It is associated with various health complications, including periodontitis and colorectal cancer. Its surface is covered with lipopolysaccharide, which interacts with the immune system and can be involved in various processes in health and disease conditions. Here we present the results of structural analysis of core oligosaccharides from the lipopolysaccharides of several strains of F. nucleatum. Pure compounds were isolated using mild acid hydrolysis or alkaline deacylation of the lipopolysaccharides and analyzed by NMR spectroscopy, mass-spectrometry and chemical methods. All cores analyzed had a common octasaccharide region, including five heptose residues and a non-phosphorylated 3-deoxy-d-manno-oct-2-ulosonic acid residue. The common region is substituted with different additional components specific for each strain. By structure type the F. nucleatum core is similar to that produced by Aeromonas.
13C NMR, 1H NMR, NMR-2D, 31P NMR, ESI-MS, mild acid hydrolysis, HPAEC, chemical methods, deamination, GPC, alkaline deacylation, fermentation
deacylated product with a part of PS repeating unit. AcOH released core additionally contained PCho at C6 of aDGlcpN6P residue; NMR temperature: 298 or 313.
13C NMR data:
Linkage Residue C1 C2 C3 C4 C5 C6 C7 C8
0,6,6,5,3,2,7 aDGlcpN 96.4 55.2 72.8 70.4 74.2 61.5
0,6,6,5,3,2 aXLLmanHepp 102.8 72.0 71.8 67.4 73.7 70.1 71.2
0,6,6,5,3 aXLLmanHepp 99.3 81.4 71.2 67.6 72.7 69.5 64.4
0,6,6,5,4,4,4,6,3 aDGlcp 99.0 71.6 73.5 70.5 74.6 61.9
0,6,6,5,4,4,4,6 bDFucpN4N 100.9 52.6 75.1 52.7 69.0 16.7
0,6,6,5,4,4,4 aDGlcpN 97.0 55.2 70.4 71.1 72.3 70.2
0,6,6,5,4,4,6 P
0,6,6,5,4,4 bDGalp 104.6 71.4 72.8 77.4 73.6 64.1
0,6,6,5,4,6,6,6,0 %xXCho
0,6,6,5,4,6,6,6 P
0,6,6,5,4,6,6 aDGlcpN 96.2 55.4 70.5 70.3 73.0 64.9
0,6,6,5,4,6 aXLLmanHepp 102.2 70.6 72.5 67.4 ? 76.7 63.5
0,6,6,5,4 bDGlcpN 99.7 58.1 72.5 77.8 75.8 66.4
0,6,6,5,6 aXLLmanHepp 101.3 71.8 71.3 67.5 73.6 70.6 64.3
0,6,6,5,7 P
0,6,6,5 aXLLmanHepp 101.6 70.8 73.2 75.1 72.0 77.9 65.9
0,6,6 aXKdop ? ? 36.2 66.4 75.7 72.9 70.5 64.8
0,6,4 P
0,6 bDGlcpN 100.2 56.6 75.3 72.8 75.0 63.5
0 aDGlcpN 92.7 55.3 70.7 71.0 73.6 70.2
P
1H NMR data:
Linkage Residue H1 H2 H3 H4 H5 H6 H7 H8
0,6,6,5,3,2,7 aDGlcpN 5.18 3.39 3.91 3.60 3.68 3.83-3.89
0,6,6,5,3,2 aXLLmanHepp 5.15 4.03 3.88 3.80 ? 4.28 3.60-4.00
0,6,6,5,3 aXLLmanHepp 5.70 3.96 3.97 3.91 3.67 4.12 3.68-3.78
0,6,6,5,4,4,4,6,3 aDGlcp 5.19 3.66 3.83 3.43 3.88 3.76-3.92
0,6,6,5,4,4,4,6 bDFucpN4N 4.78 3.37 4.41 3.94 4.11 1.39
0,6,6,5,4,4,4 aDGlcpN 5.39 3.36 3.92 3.50 4.58 3.90-4.26
0,6,6,5,4,4,6 P
0,6,6,5,4,4 bDGalp 4.53 3.63 3.91 4.21 3.85 3.98-4.05
0,6,6,5,4,6,6,6,0 %xXCho
0,6,6,5,4,6,6,6 P
0,6,6,5,4,6,6 aDGlcpN 5.59 3.44 4.00 3.63 4.00 4.15-4.15
0,6,6,5,4,6 aXLLmanHepp 5.09 4.00 3.72 3.96 3.72 4.29 3.94-3.99
0,6,6,5,4 bDGlcpN 4.88 3.25 3.71 3.84 3.64 4.00-4.12
0,6,6,5,6 aXLLmanHepp 5.37 4.09 3.90 3.86 3.81 4.05 3.74-3.90
0,6,6,5,7 P
0,6,6,5 aXLLmanHepp 5.14 4.17 4.06 4.40 4.39 4.17 4.19-4.19
0,6,6 aXKdop - - 1.81-2.10 4.25 4.17 3.77 3.76 3.70-3.95
0,6,4 P
0,6 bDGlcpN 4.90 3.14 3.94 3.91 3.77 3.56-3.67
0 aDGlcpN 5.73 3.42 3.93 3.52 4.14 3.91-4.26
P
1H/13C HSQC data:
Linkage Residue C1/H1 C2/H2 C3/H3 C4/H4 C5/H5 C6/H6 C7/H7 C8/H8
0,6,6,5,3,2,7 aDGlcpN 96.4/5.18 55.2/3.39 72.8/3.91 70.4/3.60 74.2/3.68 61.5/3.83-3.89
0,6,6,5,3,2 aXLLmanHepp 102.8/5.15 72.0/4.03 71.8/3.88 67.4/3.80 73.7/? 70.1/4.28 71.2/3.60-4.00
0,6,6,5,3 aXLLmanHepp 99.3/5.70 81.4/3.96 71.2/3.97 67.6/3.91 72.7/3.67 69.5/4.12 64.4/3.68-3.78
0,6,6,5,4,4,4,6,3 aDGlcp 99.0/5.19 71.6/3.66 73.5/3.83 70.5/3.43 74.6/3.88 61.9/3.76-3.92
0,6,6,5,4,4,4,6 bDFucpN4N 100.9/4.78 52.6/3.37 75.1/4.41 52.7/3.94 69.0/4.11 16.7/1.39
0,6,6,5,4,4,4 aDGlcpN 97.0/5.39 55.2/3.36 70.4/3.92 71.1/3.50 72.3/4.58 70.2/3.90-4.26
0,6,6,5,4,4,6 P
0,6,6,5,4,4 bDGalp 104.6/4.53 71.4/3.63 72.8/3.91 77.4/4.21 73.6/3.85 64.1/3.98-4.05
0,6,6,5,4,6,6,6,0 %xXCho
0,6,6,5,4,6,6,6 P
0,6,6,5,4,6,6 aDGlcpN 96.2/5.59 55.4/3.44 70.5/4.00 70.3/3.63 73.0/4.00 64.9/4.15-4.15
0,6,6,5,4,6 aXLLmanHepp 102.2/5.09 70.6/4.00 72.5/3.72 67.4/3.96 ?/3.72 76.7/4.29 63.5/3.94-3.99
0,6,6,5,4 bDGlcpN 99.7/4.88 58.1/3.25 72.5/3.71 77.8/3.84 75.8/3.64 66.4/4.00-4.12
0,6,6,5,6 aXLLmanHepp 101.3/5.37 71.8/4.09 71.3/3.90 67.5/3.86 73.6/3.81 70.6/4.05 64.3/3.74-3.90
0,6,6,5,7 P
0,6,6,5 aXLLmanHepp 101.6/5.14 70.8/4.17 73.2/4.06 75.1/4.40 72.0/4.39 77.9/4.17 65.9/4.19-4.19
0,6,6 aXKdop 36.2/1.81-2.10 66.4/4.25 75.7/4.17 72.9/3.77 70.5/3.76 64.8/3.70-3.95
0,6,4 P
0,6 bDGlcpN 100.2/4.90 56.6/3.14 75.3/3.94 72.8/3.91 75.0/3.77 63.5/3.56-3.67
0 aDGlcpN 92.7/5.73 55.3/3.42 70.7/3.93 71.0/3.52 73.6/4.14 70.2/3.91-4.26
P