Extracellular production of recombinant N-glycosylated anti-VEGFR2 monobody in leaky Escherichia coli strain
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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.
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.
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.
KeywordsExtracellular production Leaky Escherichia coli Monobody N-glycosylation
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.
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