Advertisement

Biochemistry (Moscow)

, Volume 80, Issue 8, pp 1073–1079 | Cite as

Asp141 and the hydrogen-bond chain Asp141–Asn109–Asp33 are respectively essential for GT80 sialyltransferase activity and structural stability

  • Xiaoyan Chen
  • Yuanming Wang
  • Zhenping Ma
  • Na Li
  • Weiqing Han
  • Qi Zhang
  • Yumei CaiEmail author
  • Jiansong ChengEmail author
Article

Abstract

Sialyltransferases are key enzymes involved in the biosynthesis of biologically and pathologically important sialic acid-containing molecules in nature. In this study, the activity of a putative sialyltransferase (Pm0160) harboring an inherent mutation D141Y in the conserved DDG motif, which has been identified in GT52 and GT80 families, was restored by reverse mutation. More interestingly, a hydrogen-bond chain was found to form between three conserved residues (Asp141, Asn109, and Asp33) of GT80 sialyltransferases based on recently determined crystal structures. Our mutagenesis experiments demonstrated that the hydrogen-bond chain connecting the general base Asp141 with Nβ4, Nβ1, and Nα1 plays an essential role in maintaining protein structural stability other than keeping the general base Asp141 in a productive orientation for sialic acid transfer.

Key words

sialyltransferase hydrogen-bond chain general base stability activity 

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. 1.
    Chen, X., and Varki, A. (2010) Advances in the biology and chemistry of sialic acids, ACS Chem. Biol., 5, 163–176.PubMedCentralCrossRefPubMedGoogle Scholar
  2. 2.
    Varki, A. (2008) Sialic acids in human health and disease, Trends Mol. Med., 14, 351–360.PubMedCentralCrossRefPubMedGoogle Scholar
  3. 3.
    Yu, H., Chokhawala, H. A., Huang, S., and Chen, X. (2006) One-pot three-enzyme chemoenzymatic approach to the synthesis of sialosides containing natural and nonnatural functionalities, Nat. Protoc., 1, 2485–2492.PubMedCentralCrossRefPubMedGoogle Scholar
  4. 4.
    Yamamoto, T. (2010) Marine bacterial sialyltransferases, Mar. Drugs, 8, 2781–2794.PubMedCentralCrossRefPubMedGoogle Scholar
  5. 5.
    Jeanneau, C., Chazalet, V., Auge, C., Soumpasis, D. M., Harduin-Lepers, A., Delannoy, P., Imberty, A., and Breton, C. (2004) Structure-function analysis of the human sialyltransferase ST3Gal I: role of n-glycosylation and a novel conserved sialyl motif, J. Biol. Chem., 279, 13461–13468.CrossRefPubMedGoogle Scholar
  6. 6.
    Datta, A. K. (2009) Comparative sequence analysis in the sialyltransferase protein family: analysis of motifs, Curr. Drug Targets, 10, 483–498.CrossRefPubMedGoogle Scholar
  7. 7.
    Freiberger, F., Claus, H., Gunzel, A., Oltmann-Norden, I., Vionnet, J., Muhlenhoff, M., Vogel, U., Vann, W. F., Gerardy-Schahn, R., and Stummeyer, K. (2007) Biochemical characterization of a Neisseria meningitidis polysialyltransferase reveals novel functional motifs in bacterial sialyltransferases, Mol. Microbiol., 65, 1258–1275.PubMedCentralCrossRefPubMedGoogle Scholar
  8. 8.
    Lin, L. Y., Rakic, B., Chiu, C. P., Lameignere, E., Wakarchuk, W. W., Withers, S. G., and Strynadka, N. C. (2011) Structure and mechanism of the lipooligosaccharide sialyltransferase from Neisseria meningitidis, J. Biol. Chem., 286, 37237–37248.Google Scholar
  9. 9.
    Ni, L., Chokhawala, H. A., Cao, H., Henning, R., Ng, L., Huang, S., Yu, H., Chen, X., and Fisher, A. J. (2007) Crystal structures of Pasteurella multocida sialyltransferase complexes with acceptor and donor analogues reveal substrate binding sites and catalytic mechanism, Biochemistry, 46, 6288–6298.CrossRefPubMedGoogle Scholar
  10. 10.
    Kim, D. U., Yoo, J. H., Lee, Y. J., Kim, K. S., and Cho, H. S. (2008) Structural analysis of sialyltransferase PM0188 from Pasteurella multocida complexed with donor analogue and acceptor sugar, BMB Rep., 41, 48–54.CrossRefPubMedGoogle Scholar
  11. 11.
    Iwatani, T., Okino, N., Sakakura, M., Kajiwara, H., Takakura, Y., Kimura, M., Ito, M., Yamamoto, T., and Kakuta, Y. (2009) Crystal structure of alpha/beta-galactoside alpha2,3-sialyltransferase from a luminous marine bacterium Photobacterium phosphoreum, FEBS Lett., 583, 2083–2087.CrossRefPubMedGoogle Scholar
  12. 12.
    Kakuta, Y., Okino, N., Kajiwara, H., Ichikawa, M., Takakura, Y., Ito, M., and Yamamoto, T. (2008) Crystal structure of Vibrionaceae Photobacterium sp. JT-ISH-224 alpha2,6-sialyltransferase in a ternary complex with donor product CMP and acceptor substrate lactose: catalytic mechanism and substrate recognition, Glycobiology, 18, 66–73.CrossRefPubMedGoogle Scholar
  13. 13.
    Lainson, F. A., Dagleish, M. P., Fontaine, M. C., Bayne, C., and Hodgson, J. C. (2013) Draft genome sequence of Pasteurella multocida A:3 strain 671/90, Genome Announc., 1, DOI:  10.1128/genomeA.00803-13.Google Scholar
  14. 14.
    Yu, H., Chokhawala, H., Karpel, R., Yu, H., Wu, B., Zhang, J., Zhang, Y., Jia, Q., and Chen, X. (2005) A multifunctional Pasteurella multocida sialyltransferase: a powerful tool for the synthesis of sialoside libraries, J. Am. Chem. Soc., 127, 17618–17619.Google Scholar
  15. 15.
    Yu, H., Cheng, J., Ding, L., Khedri, Z., Chen, Y., Chin, S., Lau, K., Tiwari, V. K., and Chen, X. (2009) Chemoenzymatic synthesis of GD3 oligosaccharides and other disialyl glycans containing natural and non-natural sialic acids, J. Am. Chem. Soc., 131, 18467–18477.Google Scholar
  16. 16.
    Sugiarto, G., Lau, K., Li, Y., Khedri, Z., Yu, H., Le, D. T., and Chen, X. (2011) Decreasing the sialidase activity of multifunctional Pasteurella multocida α2-3-sialyltransferase 1 (PmST1) by site-directed mutagenesis, Mol. Biosyst., 7, 3021–3027.CrossRefPubMedGoogle Scholar

Copyright information

© Pleiades Publishing, Ltd. 2015

Authors and Affiliations

  • Xiaoyan Chen
    • 1
  • Yuanming Wang
    • 2
  • Zhenping Ma
    • 2
  • Na Li
    • 2
  • Weiqing Han
    • 2
  • Qi Zhang
    • 2
  • Yumei Cai
    • 1
    Email author
  • Jiansong Cheng
    • 2
    Email author
  1. 1.College of Animal Science and Veterinary MedicineShandong Agricultural UniversityShandongChina
  2. 2.State Key Laboratory of Medicinal Chemical Biology and College of PharmacyNankai UniversityTianjinChina

Personalised recommendations