| Technique(s) used for Glycosylation Detection | Biotin-hydrazide labeling after periodate oxidation, sialic acid-specific lectin LFA (limax flavus agglutinin)-binding, mass excess (approx. 6 kDa) detected by intact mass analysis using ESMS (electrospray mass spectrometry) |
| Technique(s) used for Glycosylated Residue(s) Detection | Edman sequencing |
| Protein Glycosylation- Implication | Some level of glycosylation is necessary for filament assembly. Modification with either Pse5Ac7Ac or PseAm is sufficient for filament assembly. Modification with pseudaminic acid and derivatives is essential for targeting and/or secretion of flagellin. Glycosylation may be required for interaction of the subunits with the flagellin chaperone, FliS, with other components of the flagellar structure or in subunit interactions. |
| Glycan Information |
| Glycan Annotation | More Pse5Ac7Ac (5,7-diacetamido-3,5,7,9-tetradeoxy-L-glycero-L-manno-nonulosonic acid) in this case than from 81-176. Acetamidino and N-methylacetimidoyl derivatives of legionaminic acid, 315 Da Leg5Am7Ac (5-acetamidino-7-acetamido-3,5,7,9,-tetradeoxy-D-glycero-D-galacto-nonulosonic acid) and Leg5AmNMe7Ac (5-E/Z-N-(N-methylacetimidoyl)-7-acetamidino-3,5,7,9-tetradeoxy-D-glycero-D-galacto-nonulosonic), are also present. Leg5Ac7Ac (Leg) is legionaminic acid and Pse5Ac7Ac (Pse) is pseudaminic acid. |
| BCSDB ID | 21877 |
| GlyTouCan | G07086CL |
| Technique(s) used for Glycan Identification | Homo- and heteronuclear correlated two-dimensional 1H NMR, 13C HSQC (heteronuclear single quantum coherence), HMBC (heteronuclear multiple bond coherence), COSY (correlated spectroscopy), TOCSY (total correlation spectroscopy), and NOESY (nuclear Overhauser effect spectroscopy). |
| Protein Glycosylation linked (PGL) gene(s) |
| Characterized Accessory Gene(s) | Products of ptmA-H genes are involved in the biosynthesis of Leg and Pse sugars that modify the flagellin. |
| Predicted Accessory Gene(s) | ptmC, ptmB, ptmG, and ptmH may encode a Leg5Ac7Ac synthase, a CMP-Leg5Ac7Ac synthetase, a CMP-Leg5Am7Ac acetamidino synthase, and an acetamidino N-methyltransferase, respectively. These genes have been shown to be involved in the Leg biosynthesis pathway. |
| Additional Comment | It is the first report of legionaminic acid in Campylobacter sp. and the first report of legionaminic acid derivatives as modifications on a protein.
Sequon feature: Consensus sequon does not appear to be related to the site of glycosylation. However, it has been suggested that the local hydrophobicity upstream of Ser/Thr residues partially influences the site of glycosylation.
It is the first report of eubacterial flagellin glycosylation. The modified residues were identified in 1989 using Edman sequencing (Ref. no. 8) but were not known to be glycosylated by then. Glycosyl residues on some of these sites were first identified in 2001 (Ref. no. 5). |
| Literature |
| Year of Identification | 1989 |
| Year of Identification Month Wise | 1989.06 |
| Year of Validation | 1989 |
| Reference | Logan, S.M., Hui, J.P., Vinogradov, E., Aubry, A.J., Melanson, J.E., Kelly, J.F., Nothaft, H. and Soo, E.C., 2009. Identification of novel carbohydrate modifications on Campylobacter jejuni 11168 flagellin using metabolomics‐based approaches. The FEBS Journal, 276(4), pp.1014-1023. |
| Corresponding Author | Patricia Guerry |
| Contact | Enteric Diseases Department, Naval Medical Research Center, 503 Robert Grant Ave., Silver Spring, MD 20910, USA. |
| Reference | McNally, D.J., Aubry, A.J., Hui, J.P., Khieu, N.H., Whitfield, D., Ewing, C.P., Guerry, P., Brisson, J.R., Logan, S.M. and Soo, E.C., 2007. Targeted metabolomics analysis of Campylobacter coli VC167 reveals legionaminic acid derivatives as novel flagellar glycans. Journal of Biological Chemistry, 282(19), pp.14463-14475. |
| Corresponding Author | Susan M Logan
Evelyn C. Soo
|
| Contact | Institute for Biological Sciences, National Research Council of Canada, Ottawa, Ontario K1A 0R6, Canada.
National Research Council, Institute for Marine Biosciences, Halifax, Nova Scotia B3H 3Z1, Canada
|
| Reference | Goon, S., Kelly, J.F., Logan, S.M., Ewing, C.P. and Guerry, P., 2003. Pseudaminic acid, the major modification on Campylobacter flagellin, is synthesized via the Cj1293 gene. Molecular microbiology, 50(2), pp.659-671. |
| Corresponding Author | Patricia Guerry |
| Contact | Enteric Diseases Department, Naval Medical Research Center, 503 Robert Grant Ave., Silver Spring, MD 20910, USA. |
| Reference | Logan, S.M., Kelly, J.F., Thibault, P., Ewing, C.P. and Guerry, P., 2002. Structural heterogeneity of carbohydrate modifications affects serospecificity of Campylobacter flagellins. Molecular microbiology, 46(2), pp.587-597. |
| Corresponding Author | Patricia Guerry |
| Contact | Enteric Diseases Department, Naval Medical Research Center, 503 Robert Grant Ave., Silver Spring, MD 20910, USA. |
| Reference | Thibault, P., Logan, S.M., Kelly, J.F., Brisson, J.R., Ewing, C.P. and Guerry, P., 2001. Identification of the carbohydrate moieties and glycosylation motifs in Campylobacter jejuni flagellin. Journal of Biological Chemistry, 276(37), pp.34862-34870. |
| Corresponding Author | Susan M Logan |
| Contact | Institute for Biological Sciences, National Research Council of Canada, Ottawa, Ontario K1A 0R6, Canada. |
| Reference | Doig, P., Kinsella, N., Guerry, P. and Trust, T.J., 1996. Characterization of a post‐translational modification of Campylobacter flagellin: identification of a sero‐specific glycosyl moiety. Molecular microbiology, 19(2), pp.379-387. |
| Corresponding Author | Trevor J. Trust |
| Contact | Department of Biochemistry and Microbiology, University of Victoria, British Columbia, Canada. |
| Reference | Guerry, P., Doig, P., Alm, R.A., Burr, D.H., Kinsella, N. and Trust, T.J., 1996. Identification and characterization of genes required for post‐translational modification of Campylobacter coli VC167 flagellin. Molecular microbiology, 19(2), pp.369-378. |
| Corresponding Author | Patricia Guerry |
| Contact | Enteric Diseases Department, Naval Medical Research Center, 503 Robert Grant Ave., Silver Spring, MD 20910, USA. |
