National Research Council of Canada, Vaccine Program, Human Health Therapeutics, Ottawa Canada, K1A OR6
Cell surface polysaccharides produced by C. perfringens ATCC 13124 were analyzed using NMR, chemical and immunological methods. Two distinct polymers were identified. The more abundant PS1 had a structure based on a polymer of β-mannosamine with a number of modifications, including varying levels of substitution at O-6 with PEtN, N-acetylation, and different linkages between monosaccharides. The shortest variant of PS1 represented a lipoteichoic acid. It contained only 1-4-linkages between ManNAc residues, minor branching α-Ribf, and glucosyl-glycerol at the reducing end, which was acylated with linear saturated fatty acids C16, C18, and C20 (dominant). Other non-lipidated variants of PS1 contained less PEtN, no α-Ribf, up to 50% 1-3-linkages, and up to 25% ManN with the free amino group. The minor polysaccharide PS2 had a linear regular structure with a -4-α-Rha-3-β-Gal-4-β-GalNAc3PCho- repeating unit, where PCho indicates phosphocholine.
13C NMR, 1H NMR, NMR-2D, de-O-acylation, SDS-PAGE, sugar analysis, Western blotting, GPC, ion-exchange chromatography, HF treatment, N-acetylation, immunization, lysozyme treatment
Possible structure of polysaccharide pPS1 glycoform, the B-N-Q ([-4)b?ManpNAc6PEtN(1-]-4)b?ManNAc(1-4)b?ManpNAc6PEtN(1-) sequence was not proven, and it was not determined where and how many a-Ribf branches were present; the NMR solution was not indicated.
13C NMR data:
Linkage Residue C1 C2 C3 C4 C5 C6
?,4,4,3 a?Ribf 104.6 72.8 70.5 86.2 62.6
?,4,4,2 Ac ? 23.4
?,4,4,4,4,4,2 Ac ? 23.4
?,4,4,4,4,4,6,0 xXEtN 63.2 41.4
?,4,4,4,4,4,6 P
?,4,4,4,4,4 b?ManpN 101.0 54.3 72.8 67.4 76.2 65.2
?,4,4,4,4,2 Ac ? 23.4
?,4,4,4,4,6,0 xXEtN 63.2 41.4
?,4,4,4,4,6 P
?,4,4,4,4 b?ManpN 101.2 54.0 71.6 77.9 74.8 65.2
?,4,4,4,2 Ac ? 23.4
?,4,4,4 b?ManpN 101.2 54.0 71.6 77.9 76.1 61.3
?,4,4,6,0 xXEtN 63.2 41.4
?,4,4,6 P
?,4,4 b?ManpN 100.8 54.0 78.6 74.2 75.0 65.2
?,4 b?ManpN 98.1 55.1 69.5 76.6 76.1 61.3
? b?Glcp 103.5 73.9 74.8 78.9 76.1 61.3
x?Gro 71.9 71.7 63.6
1H NMR data:
Linkage Residue H1 H2 H3 H4 H5 H6
?,4,4,3 a?Ribf 5.29 4.10 4.00 4.02 3.66-3.71
?,4,4,2 Ac - 2.06
?,4,4,4,4,4,2 Ac - 2.06
?,4,4,4,4,4,6,0 xXEtN 4.13 3.29
?,4,4,4,4,4,6 P
?,4,4,4,4,4 b?ManpN 4.90 4.59 3.85 3.59 3.58 4.18
?,4,4,4,4,2 Ac - 2.06
?,4,4,4,4,6,0 xXEtN 4.13 3.29
?,4,4,4,4,6 P
?,4,4,4,4 b?ManpN 4.87 4.62 3.94 3.81 3.62 4.13-4.13
?,4,4,4,2 Ac - 2.06
?,4,4,4 b?ManpN 4.87 4.56 3.95 3.72 3.55 3.75-3.89
?,4,4,6,0 xXEtN 4.13 3.29
?,4,4,6 P
?,4,4 b?ManpN 4.87 4.72 4.00 4.05 3.63 4.13-4.13
?,4 b?ManpN 4.99 3.84 4.06 3.79 3.55 3.75-3.89
? b?Glcp 4.49 3.35 3.68 3.74 3.59 3.75-3.89
x?Gro 3.76-3.91 3.93 3.56-3.64
1H/13C HSQC data:
Linkage Residue C1/H1 C2/H2 C3/H3 C4/H4 C5/H5 C6/H6
?,4,4,3 a?Ribf 104.6/5.29 72.8/4.10 70.5/4.00 86.2/4.02 62.6/3.66-3.71
?,4,4,2 Ac 23.4/2.06
?,4,4,4,4,4,2 Ac 23.4/2.06
?,4,4,4,4,4,6,0 xXEtN 63.2/4.13 41.4/3.29
?,4,4,4,4,4,6 P
?,4,4,4,4,4 b?ManpN 101.0/4.90 54.3/4.59 72.8/3.85 67.4/3.59 76.2/3.58 65.2/4.18
?,4,4,4,4,2 Ac 23.4/2.06
?,4,4,4,4,6,0 xXEtN 63.2/4.13 41.4/3.29
?,4,4,4,4,6 P
?,4,4,4,4 b?ManpN 101.2/4.87 54.0/4.62 71.6/3.94 77.9/3.81 74.8/3.62 65.2/4.13-4.13
?,4,4,4,2 Ac 23.4/2.06
?,4,4,4 b?ManpN 101.2/4.87 54.0/4.56 71.6/3.95 77.9/3.72 76.1/3.55 61.3/3.75-3.89
?,4,4,6,0 xXEtN 63.2/4.13 41.4/3.29
?,4,4,6 P
?,4,4 b?ManpN 100.8/4.87 54.0/4.72 78.6/4.00 74.2/4.05 75.0/3.63 65.2/4.13-4.13
?,4 b?ManpN 98.1/4.99 55.1/3.84 69.5/4.06 76.6/3.79 76.1/3.55 61.3/3.75-3.89
? b?Glcp 103.5/4.49 73.9/3.35 74.8/3.68 78.9/3.74 76.1/3.59 61.3/3.75-3.89
x?Gro 71.9/3.76-3.91 71.7/3.93 63.6/3.56-3.64
report error
Found 1 record.
Displayed record 1
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nrc-cnrc.gc.ca