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Applied Microbiology and Biotechnology

, Volume 101, Issue 7, pp 2843–2851 | Cite as

Expression and secretion of glycosylated heparin biosynthetic enzymes using Komagataella pastoris

  • Jacob A. Englaender
  • Yuanyuan Zhu
  • Abhijit N. Shirke
  • Lei Lin
  • Xinyue Liu
  • Fuming Zhang
  • Richard A. Gross
  • Mattheos A. G. KoffasEmail author
  • Robert J. LinhardtEmail author
Biotechnologically relevant enzymes and proteins

Abstract

Heparin, an anticoagulant drug, is biosynthesized in selected animal cells. The heparin biosynthetic enzymes mainly consist of sulfotransferases and all are integral transmembrane glycoproteins. These enzymes are generally produced in engineered Escherichia coli as without their transmembrane domains as non-glycosylated fusion proteins. In this study, we used the yeast, Komagataella pastoris, to prepare four sulfotransferases involved in heparin biosynthesis as glycoproteins. While the yields of these yeast-expressed enzymes were considerably lower than E. coli-expressed enzymes, these enzymes were secreted into the fermentation media simplifying their purification and were endotoxin free. The activities of these sulfotransferases, expressed as glycoproteins in yeast, were compared to the bacterially expressed proteins. The yeast-expressed sulfotransferase glycoproteins showed improved kinetic properties than the bacterially expressed proteins.

Keywords

Heparin Biosynthetic enzymes Yeast expression Bacterial expression Kinetics 

Notes

Acknowledgements

The authors gratefully acknowledge funding from the National Institutes of Health (HL096972), the National Science Foundation (MCB-1448657), and funding from the China Scholarship Council.

Compliance with ethical standards

This article does not contain any studies with human participants or animals performed by any of the authors.

Conflict of interest

The authors declare that they have no conflict of interest.

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Copyright information

© Springer-Verlag Berlin Heidelberg 2016

Authors and Affiliations

  • Jacob A. Englaender
    • 1
  • Yuanyuan Zhu
    • 2
  • Abhijit N. Shirke
    • 3
  • Lei Lin
    • 3
  • Xinyue Liu
    • 3
  • Fuming Zhang
    • 4
  • Richard A. Gross
    • 3
  • Mattheos A. G. Koffas
    • 1
    • 4
    Email author
  • Robert J. Linhardt
    • 1
    • 3
    • 4
    Email author
  1. 1.Department of BiologyRensselaer Polytechnic InstituteTroyUSA
  2. 2.Department of Chemical Processing Engineering of Forest ProductsNanjing Forestry UniversityNanjingChina
  3. 3.Chemistry and Chemical BiologyRensselaer Polytechnic InstituteTroyUSA
  4. 4.Chemical and Biological Engineering and Biomedical Engineering, Center for Biotechnology and Interdisciplinary StudiesRensselaer Polytechnic InstituteTroyUSA

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