Production of Sialic Acid and Its Derivatives by Metabolic Engineering of Escherichia coli

  • Baixue Lin
  • Yong TaoEmail author
Part of the Green Chemistry and Sustainable Technology book series (GCST)


Sialic acid and its derivatives have attracted considerable interest due to their important biological functions, and are valuable resources with increasing demand in many fields. Significant efforts have been made for the efficient production of sialic acids. In this chapter, we review the achievements that have been obtained in the development of microbial processes for the production of sialic acids and its derivatives with an emphasis on the strategies of metabolic engineering for the enhanced production of N-acetylneuraminic acid, polysialic acid, and sialylated oligosaccharides, such as 3′ sialyllactose.


Metabolic engineering Escherichia coli N-Acetylneuraminic acid Polysialic acid 3′ sialyllactose 


Conflict of interest

There is no conflict of interest.


  1. 1.
    Schauer R (2004) Sialic acids: fascinating sugars in higher animals and man. Zoology 107(1):49–64CrossRefGoogle Scholar
  2. 2.
    Almagro-Moreno S, Boyd EF (2009) Insights into the evolution of sialic acid catabolism among bacteria. BMC Evol Biol 9(1):118CrossRefGoogle Scholar
  3. 3.
    Chen X, Varki A (2010) Advances in the biology and chemistry of sialic acids. ACS Chem Biol 5(2):163–176CrossRefGoogle Scholar
  4. 4.
    Schauer R (2000) Achievements and challenges of sialic acid research. Glycoconj J 17(7):485–499CrossRefGoogle Scholar
  5. 5.
    Varki NM, Varki A (2007) Diversity in cell surface sialic acid presentations: implications for biology and disease. Lab Invest 87(9):851–857CrossRefGoogle Scholar
  6. 6.
    Maru I, Ohnishi J, Ohta Y, Tsukada Y (2002) Why is sialic acid attracting interest now? Complete enzymatic synthesis of sialic acid with N-acylglucosamine 2-epimerase. J Biosci Bioeng 93(3):258–265CrossRefGoogle Scholar
  7. 7.
    Lundgren BR, Boddy CN (2007) Sialic acid and N-acyl sialic acid analog production by fermentation of metabolically and genetically engineered Escherichia coli. Org Biomol Chem 5(12):1903–1909CrossRefGoogle Scholar
  8. 8.
    Tanner ME (2005) The enzymes of sialic acid biosynthesis. Bioorg Chem 33(3):216–228CrossRefGoogle Scholar
  9. 9.
    Wu W, Air GM (2004) Binding of influenza viruses to sialic acids: reassortant viruses with A/NWS/33 hemagglutinin bind to α2, 8-linked sialic acid. Virology 325(2):340–350CrossRefGoogle Scholar
  10. 10.
    Kawai N, Ikematsu H, Iwaki N, Maeda T, Kawashima T, Hirotsu N, Kashiwagi S (2009) Comparison of the effectiveness of zanamivir and oseltamivir against influenza A/H1N1, A/H3N2, and B. Clin Infect Dis 48(7):996–997CrossRefGoogle Scholar
  11. 11.
    von Itzstein M, Dyason JC, Oliver SW, White HF, Wu W-Y, Kok GB, Pegg MS (1996) A study of the active site of influenza virus sialidase: an approach to the rational design of novel anti-influenza drugs. J Med Chem 39(2):388–391CrossRefGoogle Scholar
  12. 12.
    Zhang X, Kiechle FL (2004) Glycosphingolipids in health and disease. Ann Clin Lab Sci 34(1):3–13Google Scholar
  13. 13.
    Sprenger N, Julita M, Donnicola D, Jann A (2009) Sialic acid feeding aged rats rejuvenates stimulated salivation and colon enteric neuron chemotypes. Glycobiology 19(12):1492–1502CrossRefGoogle Scholar
  14. 14.
    Wang B, McVeagh P, Petocz P, Brand-Miller J (2003) Brain ganglioside and glycoprotein sialic acid in breastfed compared with formula-fed infants. Am J Clin Nutr 78(5):1024–1029Google Scholar
  15. 15.
    Wang B, Yu B, Karim M, Hu H, Sun Y, McGreevy P, Petocz P, Held S, Brand-Miller J (2007) Dietary sialic acid supplementation improves learning and memory in piglets. Am J Clin Nutr 85(2):561–569Google Scholar
  16. 16.
    Steinhaus S, Stark Y, Bruns S, Haile Y, Scheper T, Grothe C, Behrens P (2010) Polysialic acid immobilized on silanized glass surfaces: a test case for its use as a biomaterial for nerve regeneration. J Mater Sci Mater Med 21(4):1371–1378CrossRefGoogle Scholar
  17. 17.
    Sprenger N, Duncan PI (2012) Sialic acid utilization. Adv Nutr Int Rev J 3(3):392S–397SCrossRefGoogle Scholar
  18. 18.
    Bode L (2006) Recent advances on structure, metabolism, and function of human milk oligosaccharides. J Nutr 136(8):2127–2130Google Scholar
  19. 19.
    Martin JE, Tanenbaum SW, Flashner M (1977) A facile procedure for the isolation of N-acetylneuramic acid from edible bird’s-nest. Carbohydr Res 56(2):423CrossRefGoogle Scholar
  20. 20.
    Koketsu M, Juneja LR, Kawanami H, Kim M, Yamamoto T (1992) Preparation of N-acetylneuraminic acid from delipidated egg yolk. Glycoconj J 9(2):70–74CrossRefGoogle Scholar
  21. 21.
    Ferrero MA, Reglero A, Fernandez-Lopez M, Ordas R, Rodriguez-Aparicio LB (1996) N-acetyl-d-neuraminic acid lyase generates the sialic acid for colominic acid biosynthesis in Escherichia coli K1. Biochem J 317(Pt 1):157–165CrossRefGoogle Scholar
  22. 22.
    Rodriguez-Aparicio L, Reglero A, Ortiz A, Luengo J (1988) Effect of physical and chemical conditions on the production of colominic acid by Escherichia coli in a defined medium. Appl Microbiol Biotechnol 27(5):474–483CrossRefGoogle Scholar
  23. 23.
    Zhan X, Zhu L, Wu J, Zhen Z, Jia W (2002) Production of polysialic acid from fed-batch fermentation with pH control. Biochem Eng J 11(2):201–204CrossRefGoogle Scholar
  24. 24.
    Wang TH, Chen YY, Pan HH, Wang FP, Cheng CH, Lee WC (2009) Production of N-acetyl-d-neuraminic acid using two sequential enzymes overexpressed as double-tagged fusion proteins. BMC Biotechnol 9:63CrossRefGoogle Scholar
  25. 25.
    Hu S, Chen J, Yang Z, Shao L, Bai H, Luo J, Jiang W, Yang Y (2010) Coupled bioconversion for preparation of N-acetyl-d-neuraminic acid using immobilized N-acetyl-d-glucosamine-2-epimerase and N-acetyl-d-neuraminic acid lyase. Appl Microbiol Biotechnol 85(5):1383–1391CrossRefGoogle Scholar
  26. 26.
    Vimr ER, Kalivoda KA, Deszo EL, Steenbergen SM (2004) Diversity of microbial sialic acid metabolism. Microbiol Mol Biol Rev 68(1):132–153CrossRefGoogle Scholar
  27. 27.
    Martinez J, Steenbergen S, Vimr E (1995) Derived structure of the putative sialic acid transporter from Escherichia coli predicts a novel sugar permease domain. J Bacteriol 177(20):6005–6010CrossRefGoogle Scholar
  28. 28.
    Ferrero MÁ, Aparicio LR (2010) Biosynthesis and production of polysialic acids in bacteria. Appl Microbiol Biotechnol 86(6):1621–1635CrossRefGoogle Scholar
  29. 29.
    Kalivoda KA, Steenbergen SM, Vimr ER, Plumbridge J (2003) Regulation of sialic acid catabolism by the DNA binding protein NanR in Escherichia coli. J Bacteriol 185(16):4806–4815CrossRefGoogle Scholar
  30. 30.
    Tao F, Zhang Y, Ma C, Xu P (2011) One-pot bio-synthesis: N-acetyl-d-neuraminic acid production by a powerful engineered whole-cell catalyst. Sci Rep 1:1–7CrossRefGoogle Scholar
  31. 31.
    Boddy CN, Lundgren BR (2009) Metabolically engineered Escherichia coli for enhanced production of sialic acid. US Patent (PCT/US2007/079496)Google Scholar
  32. 32.
    Ishikawa M, Koizumi S (2010) Microbial production of N-acetylneuraminic acid by genetically engineered Escherichia coli. Carbohydr Res 345(18):2605–2609CrossRefGoogle Scholar
  33. 33.
    Kang J, Gu P, Wang Y, Li Y, Yang F, Wang Q, Qi Q (2012) Engineering of an N-acetylneuraminic acid synthetic pathway in Escherichia coli. Metab Eng 14(6):623–629CrossRefGoogle Scholar
  34. 34.
    Lin B-X, Zhang Z-J, Liu W-F, Dong Z-Y, Tao Y (2013) Enhanced production of N-acetyl-d-neuraminic acid by multi-approach whole-cell biocatalyst. Appl Microbiol Biotechnol 97(11):4775–4784CrossRefGoogle Scholar
  35. 35.
    Lee YC, Chien HC, Hsu WH (2007) Production of N-acetyl-d-neuraminic acid by recombinant whole cells expressing Anabaena sp. CH1 N-acetyl-d-glucosamine 2-epimerase and Escherichia coli N-acetyl-d-neuraminic acid lyase. J Biotechnol 129(3):453–460CrossRefGoogle Scholar
  36. 36.
    Tabata K, Koizumi S, Endo T, Ozaki A (2002) Production of N-acetyl-d-neuraminic acid by coupling bacteria expressing N-acetyl-d-glucosamine 2-epimerase and N-acetyl-d-neuraminic acid synthetase. Enzyme Microb Technol 30(3):327–333CrossRefGoogle Scholar
  37. 37.
    Zhang Y, Tao F, Du M, Ma C, Qiu J, Gu L, He X, Xu P (2010) An efficient method for N-acetyl-d-neuraminic acid production using coupled bacterial cells with a safe temperature-induced system. Appl Microbiol Biotechnol 86(2):481–489CrossRefGoogle Scholar
  38. 38.
    Tao F, Zhang Y, Ma C, Xu P (2010) Biotechnological production and applications of N-acetyl-d-neuraminic acid: current state and perspectives. Appl Microbiol Biotechnol 87(4):1281–1289CrossRefGoogle Scholar
  39. 39.
    Xu X, Gao C, Zhang X, Che B, Ma C, Qiu J, Tao F, Xu P (2011) Production of N-acetyl-d-neuraminic acid by use of an efficient spore surface display system. Appl Environ Microbiol 77(10):3197–3201CrossRefGoogle Scholar
  40. 40.
    Samain E (2008) High yield production of sialic acid (Neu5Ac) by fermentation. (WO 2008/040717)Google Scholar
  41. 41.
    Sun W, Ji W, Li N, Tong P, Cheng J, He Y, Chen Y, Chen X, Wu J, Ouyang P (2013) Construction and expression of a polycistronic plasmid encoding N-acetylglucosamine 2-epimerase and N-acetylneuraminic acid lyase simultaneously for production of N-acetylneuraminic acid. Bioresour Technol 130:23–29CrossRefGoogle Scholar
  42. 42.
    Fierfort N, Samain E (2008) Genetic engineering of Escherichia coli for the economical production of sialylated oligosaccharides. J Biotechnol 134(3):261–265CrossRefGoogle Scholar
  43. 43.
    Zimmermann V, Hennemann H-G, Daußmann T, Kragl U (2007) Modelling the reaction course of N-acetylneuraminic acid synthesis from N-acetyl-d-glucosamine—new strategies for the optimisation of neuraminic acid synthesis. Appl Microbiol Biotechnol 76(3):597–605CrossRefGoogle Scholar
  44. 44.
    Barrett B, Ebah L, Roberts I (2002) Genomic structure of capsular determinants. In: Pathogenicity islands and the evolution of pathogenic microbes. Springer, pp 137–155Google Scholar
  45. 45.
    Daines DA, Silver RP (2000) Evidence for multimerization of Neu proteins involved in polysialic acid synthesis in Escherichia coli K1 using improved LexA-based vectors. J Bacteriol 182(18):5267–5270CrossRefGoogle Scholar
  46. 46.
    Cress BF, Englaender JA, He W, Kasper D, Linhardt RJ, Koffas M (2014) Masquerading microbial pathogens: capsular polysaccharides mimic host-tissue molecules. FEMS Microbiol Rev 38:660–697CrossRefGoogle Scholar
  47. 47.
    Steenbergen SM, Lee Y-C, Vann WF, Vionnet J, Wright LF, Vimr ER (2006) Separate pathways for O acetylation of polymeric and monomeric sialic acids and identification of sialyl O-acetyl esterase in Escherichia coli K1. J Bacteriol 188(17):6195–6206CrossRefGoogle Scholar
  48. 48.
    Zapata G, Vann W, Aaronson W, Lewis M, Moos M (1989) Sequence of the cloned Escherichia coli K1 CMP-N-acetylneuraminic acid synthetase gene. J Biol Chem 264(25):14769–14774Google Scholar
  49. 49.
    Vann W, Silver R, Abeijon C, Chang K, Aaronson W, Sutton A, Finn C, Lindner W, Kotsatos M (1987) Purification, properties, and genetic location of Escherichia coli cytidine 5’-monophosphate N-acetylneuraminic acid synthetase. J Biol Chem 262(36):17556–17562Google Scholar
  50. 50.
    Liu G, Jin C, Jin C (2004) CMP-N-acetylneuraminic acid synthetase from Escherichia coli K1 is a bifunctional enzyme: identification of minimal catalytic domain for synthetase activity and novel functional domain for platelet-activating factor acetylhydrolase activity. J Biol Chem 279(17):17738–17749CrossRefGoogle Scholar
  51. 51.
    Song L, Zhou H, Cai X, Li C, Liang J, Jin C (2011) NeuA O-acetylesterase activity is specific for CMP-activated O-acetyl sialic acid in Streptococcus suis serotype 2. Biochem Biophys Res Commun 410(2):212–217CrossRefGoogle Scholar
  52. 52.
    Willis LM, Whitfield C (2013) Structure, biosynthesis, and function of bacterial capsular polysaccharides synthesized by ABC transporter-dependent pathways. Carbohydr Res 378:35–44CrossRefGoogle Scholar
  53. 53.
    Andreishcheva EN, Vann WF (2006) Gene products required for de novo synthesis of polysialic acid in Escherichia coli K1. J Bacteriol 188(5):1786–1797CrossRefGoogle Scholar
  54. 54.
    Chen F, Tao Y, Jin C, Xu Y, Lin B-X (2015) Enhanced production of polysialic acid by metabolic engineering of Escherichia coli. Appl Microbiol Biotechnol 99(6):2603–2611CrossRefGoogle Scholar
  55. 55.
    Wu J-R, Liu J-L, Zhan X-B, Lin C-C, Zhao H (2010) Enhancement of polysialic acid yield by reducing initial phosphate and feeding ammonia water to Escherichia coli CCTCC M208088. Biotechnol Bioprocess Eng 15(4):657–663CrossRefGoogle Scholar
  56. 56.
    Liu J-L, Zhan X-B, Wu J-R, Lin C-C, Yu D-F (2010) An efficient and large-scale preparation process for polysialic acid by Escherichia coli CCTCC M208088. Biochem Eng J 53(1):97–103CrossRefGoogle Scholar
  57. 57.
    Zheng Z-Y, Wang S-Z, Li G-S, Zhan X-B, Lin C-C, Wu J-R, Zhu L (2013) A new polysialic acid production process based on dual-stage pH control and fed-batch fermentation for higher yield and resulting high molecular weight product. Appl Microbiol Biotechnol 97(6):2405–2412CrossRefGoogle Scholar
  58. 58.
    Lin B-X, Qiao Y, Shi B, Tao Y (2016) Polysialic acid biosynthesis and production in Escherichia coli: current state and perspectives. Appl Microbiol Biotechnol 100:1–8CrossRefGoogle Scholar
  59. 59.
    Rode B, Endres C, Ran C, Stahl F, Beutel S, Kasper C, Galuska S, Geyer R, Muhlenhoff M, Gerardy-Schahn R, Scheper T (2008) Large-scale production and homogenous purification of long chain polysialic acids from E. coli K1. J Biotechnol 135(2):202–209CrossRefGoogle Scholar
  60. 60.
    Chen R, John J, Rode B, Hitzmann B, Gerardy-Schahn R, Kasper C, Scheper T (2011) Comparison of polysialic acid production in Escherichia coli K1 during batch cultivation and fed-batch cultivation applying two different control strategies. J Biotechnol 154(4):222–229CrossRefGoogle Scholar
  61. 61.
    Zheng ZY, Wang SZ, Li GS, Zhan XB, Lin CC, Wu JR, Zhu L (2013) A new polysialic acid production process based on dual-stage pH control and fed-batch fermentation for higher yield and resulting high molecular weight product. Appl Microbiol Biotechnol 97(6):2405–2412CrossRefGoogle Scholar
  62. 62.
    Gilbert M, Bayer R, Cunningham A-M, DeFrees S, Gao Y, Watson DC, Young NM, Wakarchuk WW (1998) The synthesis of sialylated oligosaccharides using a CMP-Neu5Ac synthetase/sialyltransferase fusion. Nat Biotechnol 16(8):769–772CrossRefGoogle Scholar
  63. 63.
    Endo T, Koizumi S, Tabata K, Ozaki A (2000) Large-scale production of CMP-NeuAc and sialylated oligosaccharides through bacterial coupling. Appl Microbiol Biotechnol 53(3):257–261CrossRefGoogle Scholar
  64. 64.
    Priem B, Gilbert M, Wakarchuk WW, Heyraud A, Samain E (2002) A new fermentation process allows large-scale production of human milk oligosaccharides by metabolically engineered bacteria. Glycobiology 12(4):235–240CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany 2016

Authors and Affiliations

  1. 1.CAS Key Laboratory of Microbial Physiological and Metabolic EngineeringInstitute of Microbiology, Chinese Academy of SciencesBeijingChina

Personalised recommendations