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

OBJECTIVE

To improve the production yield of N-glycosylated anti-VEGFR2 (vascular endothelial growth factor receptor 2) monobody (FN3-Gly) in lpp knockout Escherichia coli cells harboring Campylobacter jejuni N-glycosylation pathway.

RESULTS

The leaky CLM37-Δlpp strain efficiently secreted FN3-Gly into culture medium. The extracellular levels of glycosylated FN3-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 FN3-Gly (70 ± 3.4 mg/L) was found in culture medium via auto-induction. Furthermore, glycosylated FN3-Gly was more stable than unglycosylated FN3-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 FN3-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.