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3 Biotech

, 9:302 | Cite as

Improving production of N-glycosylated recombinant proteins by leaky Escherichia coli

  • Ning Ding
  • Yao Ruan
  • Xin Fu
  • Yue Lin
  • Hongyou Yu
  • Lichi Han
  • Changzhen Fu
  • Jianing ZhangEmail author
  • Xuejun HuEmail author
Original Article
  • 52 Downloads

Abstract

Escherichia coli has been considered as a promising host for the production of N-glycosylated therapeutic proteins and glycoconjugate vaccines. In this study, we developed a simple and efficient strategy for improving the production of N-glycosylated recombinant proteins by combining auto-induction with the use of a leaky E. coli strain. A leaky E. coli strain, designated as CLM37-Δlpp, was engineered by deleting the Braun’s lipoprotein (lpp) gene of E. coli strain CLM37. Three distinct acceptor model N-glycosylated proteins, glyco-tagged human tenth fibronectin type III domain (FN3-Gly), enhanced green fluorescent protein (eGFP-Gly), and scFv of vascular endothelial growth factor receptor 3 (scFv-VEGFR3-Gly) were then expressed in CLM37-Δlpp, which carried an N-glycosylation machinery from Campylobacter jejuni for the investigation of glycoprotein production. As much as 75%, 65%, and 60% of the glycosylated FN3-Gly, eGFP-Gly, and scFv-VEGFR3-Gly, respectively, were found in the culture medium. The yields of glycosylated FN3-Gly, eGFP-Gly, and scFv-VEGFR3-Gly were 106 ± 7.4 mg/L, 65 ± 2.5 mg/L, and 62 ± 4.3 mg/L, respectively, which were more than three folds the corresponding yields obtained when these proteins were expressed in CLM37, the unmodified strain. The results suggested that this simplified approach could improve the production of N-glycosylated proteins with E. coli to facilitate large-scale production.

Keywords

N-Glycosylation Leaky Escherichia coli Extracellular production Auto-induction 

Notes

Acknowledgements

We thank Markus Aebi and Andreas Plückthun for the kind gifts of E. coli CLM37, pACYCpgl plasmid, and pIG6 Vector. We thank Dr Alan K Chang for revising the language of the manuscript. The National Natural Science Foundation of China (31370937 to X.H.; 31570802 to J.Z.) and the Natural Science Foundation of Liaoning Province (1555548800399) funded this research.

Compliance with ethical standards

Conflict of interest

The authors declare no conflict of interest.

Supplementary material

13205_2019_1830_MOESM1_ESM.doc (349 kb)
Supplementary material 1 (DOC 349 kb)

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Copyright information

© King Abdulaziz City for Science and Technology 2019

Authors and Affiliations

  • Ning Ding
    • 1
    • 2
  • Yao Ruan
    • 2
  • Xin Fu
    • 2
  • Yue Lin
    • 2
  • Hongyou Yu
    • 2
  • Lichi Han
    • 2
  • Changzhen Fu
    • 2
  • Jianing Zhang
    • 1
    Email author
  • Xuejun Hu
    • 2
    Email author
  1. 1.School of Life Science and MedicineDalian University of TechnologyLiaoningChina
  2. 2.Academic Centre for Medical Research, Medical CollegeDalian UniversityLiaoningChina

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