Journal of Microbiology and Biotechnology

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
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Biochemical Characterization of a Glycosyltransferase Homolog from an Oral Pathogen Fusobacterium nucleatum as a Human Glycan-Modifying Enzyme

  • Kim, Seong-Hun (Omics and Integration Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB)) ;
  • Oh, Doo-Byoung (Omics and Integration Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB)) ;
  • Kwon, Oh-Suk (Omics and Integration Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB)) ;
  • Jung, Jae-Kap (Omics and Integration Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB)) ;
  • Lee, Yun-Mi (Omics and Integration Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB)) ;
  • Ko, Ki-Sung (Division of Biological Science, College of Natural Science, Wonkwang University) ;
  • Ko, Jeong-Heon (Omics and Integration Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB)) ;
  • Kang, Hyun-Ah (Omics and Integration Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB))
  • Published : 2008.05.31
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Abstract

Bacterial glycosyltransferases have drawn growing attention as economical enzymes for oligosaccharide synthesis, with their easy expression and relatively broad substrate specificity. Here, we characterized a glycosyltransferase homolog (Fnu_GT) from a human oral pathogen, Fusobacterium nucleatum. Bioinformatic analysis showed that Fnu_GT belongs to the glycosyltransferases family II. The recombinant Fnu_GT (rFnu_GT) expressed in Escherichia coli displayed the highest glycosylation activity when UDP-galactose (Gal) was used as a donor nucleotide-sugar with heptose or N-acetylglucosamine (GlcNAc) as an acceptor sugar. Interestingly, rFnu_GT transferred the galactose moiety of UDP-Gal to a nonreducing terminal GlcNAc attached to the trimannosyl core glycan, indicating its potential as an enzyme for human-type N-glycan synthesis.

Keywords

References

  1. Bachrach, G., S. K. Haake, A. Glick, R. Hazan, R. Naor, R. N. Andersen, and P. E. Kolenbrander. 2004. Characterization of the novel Fusobacterium nucleatum plasmid pKH9 and evidence of an addiction system. Appl. Environ. Microbiol. 70: 6957-6962 https://doi.org/10.1128/AEM.70.12.6957-6962.2004
  2. Bateman, A., E. Birney, R. Durbin, S. R. Eddy, K. L. Howe, and E. L. Sonnhammer. 2000. The Pfam protein families database. Nucleic Acids Res. 28: 263-266 https://doi.org/10.1093/nar/28.1.263
  3. Blixt, O., I. van Die, T. Norberg, and D. H. van den Eijnden. 1999. High-level expression of the Neisseria meningitidis lgtA gene in Escherichia coli and characterization of the encoded N-acetylglucosaminyltransferase as a useful catalyst in the synthesis of GlcNAcb1-3Gal and GalNAcb1-3Gal linkages. Glycobiology 9: 1061-1071 https://doi.org/10.1093/glycob/9.10.1061
  4. Bonofiglio, L., E. Garcia, and M. Mollerach. 2005. Biochemical characterization of the pneumococcal glucose 1-phosphate uridylyltransferase (GalU) essential for capsule biosynthesis. Curr. Microbiol. 51: 217-221 https://doi.org/10.1007/s00284-005-4466-0
  5. Charnock, S. J. and G. J. Davies. 1999. Structure of the nucleotide-diphospho-sugar transferase, SpsA from Bacillus subtilis, in native and nucleotide-complexed forms. Biochemistry 38: 6380-6385 https://doi.org/10.1021/bi990270y
  6. Coutinho, P. M., E. Deleury, G. J. Davies, and B. Henrissat. 2003. An evolving hierarchical family classification for glycosyltransferases. J. Mol. Biol. 328: 307-317 https://doi.org/10.1016/S0022-2836(03)00307-3
  7. Cox, A. D., D. W. Hood, A. Martin, K. M. Makepeace, M. E. Deadman, J. Li, J. R. Brisson, E. R. Moxon, and J. C. Richards. 2002. Identification and structural characterization of a sialylated lacto-N-neotetraose structure in the lipopolysaccharide of Haemophilus influenzae. Eur. J. Biochem. 269: 4009-4019 https://doi.org/10.1046/j.1432-1033.2002.03090.x
  8. Endo, T., S. Koizumi, K. Tabata, and A. Ozaki. 2000. Cloning and expression of $\beta$1,4-galactosyltransferase gene from Helicobacter pylori. Glycobiology 10: 809-813 https://doi.org/10.1093/glycob/10.8.809
  9. Garcia, M. M., S. A. Becker, B. W. Brooks, J. N. Berg, and S. M. Finegold. 1992. Ultrastructure and molecular characterization of Fusobacterium necrophorum biovars. Can. J. Vet. Res. 56: 318-325
  10. Garinot-Schneider, C., A. C. Lellouch, and R. A. Geremia. 2000. Identification of essential amino acid residues in the Sinorhizobium meliloti glucosyltransferase ExoM. J. Biol. Chem. 275: 31407-31413 https://doi.org/10.1074/jbc.M004524200
  11. Jewett, A., W. R. Hume, H. Le, T. N. Huynh, Y. W. Han, G.. Cheng, and W. Shi. 2000. Induction of apoptotic cell death in peripheral blood mononuclear and polymorphonuclear cells by an oral bacterium, Fusobacterium nucleatum. Infect. Immun. 68: 1893-1898 https://doi.org/10.1128/IAI.68.4.1893-1898.2000
  12. Johnson, K. F. 1999. Synthesis of oligosaccharides by bacterial enzymes. Glycoconj. J. 16: 141-146 https://doi.org/10.1023/A:1026440509859
  13. Kanipes, M. I., E. Papp-Szabo, P. Guerry, and M. A. Monteiro. 2006. Mutation of waaC, encoding heptosyltransferase I in Campylobacter jejuni 81-176, affects the structure of both lipooligosaccharide and capsular carbohydrate. J. Bacteriol. 188: 3273-3279 https://doi.org/10.1128/JB.188.9.3273-3279.2006
  14. Kapatral, V., I. Anderson, N. Ivanova, G.. Reznik, T. Los, A. Lykidis, A. Bhattacharyya, A. Bartman, W. Gardner, G. Grechkin, L. Zhu, O. Vasieva, L. Chu, Y. Kogan, O. Chaga, E. Goltsman, A. Bernal, N. Larsen, M. D'Souza, T. Walunas, G. Pusch, R. Haselkorn, M. Fonstein, N. Kyrpides, and R. Overbeek. 2002. Genome sequence and analysis of the oral bacterium Fusobacterium nucleatum strain ATCC 25586. J. Bacteriol. 184: 2005-2018 https://doi.org/10.1128/JB.184.7.2005-2018.2002
  15. Kim, J. H., B. G. Kim, J. A. Kim, Y. Park, Y. J. Lee, Y. Lim, and J.-H. Ahn. 2007. Glycosylation of flavonoids with E. coli expressing glycosyltransferase from Xanthomonas campestris. J. Microbiol. Biotechnol. 17: 539-542
  16. Lee, H. J., B. G.. Kim, and J.-H. Ahn. 2006. Molecular cloning and characterization of Bacillus cereus O-methyltransferase. J. Microbiol. Biotechnol. 16: 619-622
  17. Melaugh, W., N. J. Phillips, A. A. Campagnari, R. Karalus, and B. W. Gibson. 1992. Partial characterization of the major lipooligosaccharide from a strain of Haemophilus ducreyi, the causative agent of chancroid, a genital ulcer disease. J. Biol. Chem. 267: 13434-13439
  18. Moran, A. P., M. M. Prendergast, and B. J. Appelmelk. 1996. Molecular mimicry of host structures by bacterial lipopolysaccharides and its contribution to disease. FEMS Immunol. Med. Microbiol. 16: 105-115 https://doi.org/10.1111/j.1574-695X.1996.tb00127.x
  19. Oberthur, M., C. Leimkuhler, R. G. Kruger, W. Lu, C. T. Walsh, and D. Kahne. 2005. A systematic investigation of the synthetic utility of glycopeptide glycosyltransferase. J. Am. Chem. Soc. 127: 10747-10752 https://doi.org/10.1021/ja052945s
  20. Okuda, K., T. Kato, K. Ishihara, and Y. Naito. 1991. Adherence to experimental pellicle of rough-type lipopolysaccharides from subgingival plaque bacteria. Oral Microbiol. Immunol. 6: 241-245 https://doi.org/10.1111/j.1399-302X.1991.tb00484.x
  21. Palcic, M. M. and K. Sujino. 2001. Assays for glycosyltransferases. Trends Glycosci. Glycotechnol. 13: 361-370 https://doi.org/10.4052/tigg.13.361
  22. Park, J. E., K. Y. Lee, S. I. Do, and S. S. Lee. 2002. Expression and characterization of $\beta$-1,4-galactosyltransferase from Neisseria meningitidis and Neisseria gonorrhoeae. J. Biochem. Mol. Biol. 35: 330-336 https://doi.org/10.5483/BMBRep.2002.35.3.330
  23. Phillips, N. J., T. J. Miller, J. J. Engstrom, W. Melaugh, R. McLaughlin, M. A. Apicella, and B. W. Gibson. 2000. Characterization of chimeric lipopolysaccharides from Escherichia coli strain JM109 transformed with lipooligosaccharide synthesis genes (lsg) from Haemophilus influenzae. J. Biol. Chem. 275: 4747-4758 https://doi.org/10.1074/jbc.275.7.4747
  24. Piekarowicz, A. and D. C. Stein. 2002. Biochemical properties of Neisseria gonorrhoeae LgtE. J. Bacteriol. 184: 6410-6416 https://doi.org/10.1128/JB.184.23.6410-6416.2002
  25. Qasba, P. K., B. Ramakrishnan, and E. Boeggeman. 2005. Substrate-induced conformational changes in glycosyltransferases. Trends Biochem. Sci. 30: 53-62 https://doi.org/10.1016/j.tibs.2004.11.005
  26. Rosen, G. and M. N. Sela. 2006. Coaggregation of Porphyromonas gingivalis and Fusobacterium nucleatum PK 1594 is mediated by capsular polysaccharide and lipopolysaccharide. FEMS Microbiol. Lett. 256: 304-310 https://doi.org/10.1111/j.1574-6968.2006.00131.x
  27. Saksouk N., L. Pelosi, P. Colin-Morel, M. Boumedienne, P. L. Abdian, and R. A. Geremia. 2005. The capsular polysaccharide biosynthesis of Streptococcus pneumoniae serotype 8: Functional identification of the glycosyltransferase WciS (Cap8H). Biochem. J. 389: 63-72 https://doi.org/10.1042/BJ20050217
  28. Shao, J., J. Zhang, P. Kowal, Y. Lu, and P. G. Wang. 2002. Overexpression and biochemical characterization of $\beta$-1,3-Nacetylgalactosaminyltransferase LgtD from Haemophilus influenzae strain Rd. Biochem. Biophys. Res. Commun. 295: 1-8 https://doi.org/10.1016/S0006-291X(02)00615-0
  29. St. Michael, F., C. M. Szymanski, J. Li, K. H. Chan, N. H. Khieu, S. Larocque, W. W. Wakarchuk, J. R. Brisson, and M. A. Monteiro. 2002. The structures of the lipooligosaccharide and capsule polysaccharide of Campylobacter jejuni genome sequenced strain NCTC 11168. Eur. J. Biochem. 269: 5119-5136 https://doi.org/10.1046/j.1432-1033.2002.03201.x
  30. Wang, Y., S. S. Wong, M. N. Fukuda, H. Zu, Z. Liu, Q. Tang, and H. E. Appert. 1994. Identification of functional cysteine residues in human galactosyltransferase. Biochem. Biophys. Res. Commun. 204: 701-709 https://doi.org/10.1006/bbrc.1994.2516
  31. Watanabe, M., K. Miyake, K. Yanae, Y. Kataoka, S. Koizumi, T. Endo, A. Ozaki, and S. Iijima. 2002. Molecular characterization of a novel $\beta$1,3-galactosyltransferase for capsular polysaccharide synthesis by Streptococcus agalactiae type Ib. J. Biochem. (Tokyo) 131: 183-191 https://doi.org/10.1093/oxfordjournals.jbchem.a003086