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Biotechnology Letters

, Volume 41, Issue 11, pp 1265–1274 | Cite as

Extracellular production of recombinant N-glycosylated anti-VEGFR2 monobody in leaky Escherichia coli strain

  • Ning Ding
  • Xin Fu
  • Yao Ruan
  • Jing Zhu
  • Pingping Guo
  • Lichi Han
  • Jianing ZhangEmail author
  • Xuejun HuEmail author
Original Research Paper
  • 104 Downloads

Abstract

Objective

To improve the production yield of N-glycosylated anti-VEGFR2 (vascular endothelial growth factor receptor 2) monobody (FN3VEGFR2-Gly) in lpp knockout Escherichia coli cells harboring Campylobacter jejuni N-glycosylation pathway.

Results

The leaky CLM37-Δlpp strain efficiently secreted FN3VEGFR2-Gly into culture medium. The extracellular levels of glycosylated FN3VEGFR2-Gly in CLM37-Δlpp culture medium were approximately 11 and 15 times higher compared to those in CLM37 cells via IPTG and auto-induction, respectively. In addition, the highest level of total glycosylated FN3VEGFR2-Gly (70 ± 3.4 mg/L) was found in culture medium via auto-induction. Furthermore, glycosylated FN3VEGFR2-Gly was more stable than unglycosylated FN3VEGFR2-Gly in this expression system, but their bioactivities were relatively similar.

Conclusions

Lpp knockout leaky E. coli strain combined with auto-induction method can enhance the extracellular production of homogenous N-glycosylated FN3VEGFR2-Gly, and facilitate the downstream protein purification. The findings of this study may provide practical implications for the large-scale production and cost-effective harvesting of N-glycosylation proteins.

Keywords

Extracellular production Leaky Escherichia coli Monobody N-glycosylation 

Notes

Acknowledgements

We thank Markus Aebi and Andreas Plückthun for the kind gifts of E. coli CLM37, pACYCpgl plasmid, and pIG6 Vector. The National Natural Science Foundation of China (NSFC Grant 31070822 and 31370937 to X.H.; NSFC Grant 31570802 to J.Z.) and the Natural Science Foundation of Liaoning Province (2019-ZD-0563) funded this research.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

10529_2019_2731_MOESM1_ESM.pdf (486 kb)
Supplementary file1 (PDF 485 kb)

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Copyright information

© Springer Nature B.V. 2019

Authors and Affiliations

  1. 1.Academic Centre for Medical Research, Medical CollegeDalian UniversityLiaoningChina
  2. 2.School of Life Science and MedicineDalian University of TechnologyLiaoningChina

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