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Engineering Sialic Acid Synthesis Ability in Insect Cells

  • Karthik Viswanathan
  • Someet Narang
  • Michael J. Betenbaugh
Part of the Methods in Molecular Biology book series (MIMB, volume 1321)

Abstract

Insect cells lack the ability to synthesize the sialic acid donor molecule CMP-sialic acid or its precursor, sialic acid. In this chapter, we describe a method to engineer CMP-sialic acid synthesis capability into Spodoptera frugiperda (Sf9) cells, a prototypical insect cell line, by recombinant expression of sialic acid synthesis pathway genes using baculovirus technology. Co-expression of a sialuria mutant UDP-GlcNAc-2-epimerase/ManNAc kinase (EKR263L), wild-type sialic acid 9-phosphate synthase (SAS), and wild-type CMP-sialic acid synthetase (CSAS) in the presence of GlcNAc leads to synthesis of CMP-sialic acids synthesis to support sialylation of N-glycans on glycoproteins.

Key words

Sialic acid Sialylation Glycosylation Sialuria N-acetylneuraminic acid Neu5Ac Insect cells Sf9 SAS CMP-SAS 

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Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  • Karthik Viswanathan
    • 1
  • Someet Narang
    • 2
  • Michael J. Betenbaugh
    • 3
  1. 1.Department of Biological EngineeringMassachusetts Institute of TechnologyCambridgeUSA
  2. 2.Cell Culture and Fermentation SciencesGaithersburgUSA
  3. 3.Department of Chemical and Biomolecular EngineeringJohns Hopkins UniversityBaltimoreUSA

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