Preprint Article Version 1 Preserved in Portico This version is not peer-reviewed

Biological Activity of Optimized Codon Bovine Type III Interferon Expressed in Pichia Pastoris

Ran An
,
Runxiang Zhang
ORCID logo ,
Yongli Guo
,
Jinfeng Geng
,
Minglu Si
,
Shuangfeng Wang
,
Mingchun Gao
* ,
Junwei Wang
*
Version 1 : Received: 16 April 2023 / Approved: 17 April 2023 / Online: 17 April 2023 (10:53:42 CEST)

A peer-reviewed article of this Preprint also exists.

An, R.; Zhang, R.; Guo, Y.; Geng, J.; Si, M.; Wang, S.; Gao, M.; Wang, J. Biological Activity of Optimized Codon Bovine Type III Interferon Expressed in Pichia pastoris. Viruses 2023, 15, 1101. An, R.; Zhang, R.; Guo, Y.; Geng, J.; Si, M.; Wang, S.; Gao, M.; Wang, J. Biological Activity of Optimized Codon Bovine Type III Interferon Expressed in Pichia pastoris. Viruses 2023, 15, 1101.

Abstract

Type III interferon (IFN-λs) exhibit potent antiviral activity and immunomodulatory effects in specific cells. Nucleotide fragements of bovine ifn-λ (boifn-λ) gene was synthetized after codon optimization. The boifn-λ gene was then amplified by splicing by overlap extension PCR (SOE PCR), resulting in serendipitous acquisition of the mutated boIFN-λ3V18M. Recombinant plasmids pPICZαA-boIFN-λ3/λ3V18M were constructed and the corresponding proteins were expressed in Pichia pastoris with high-level extracellular soluble form. Dominant expression strains of boIFN-λ3/λ3V18M were selected by Western blot and ELISA, cultured on a large scale, and the recombinant proteins purified by ammonium sulphate precipitation and ion exchange chromatography, yielding 1.5g/L and 0.3 g/L, with 85% and 92% purity, respectively. The antiviral activities of boIFN-λ3/λ3V18M exceeded 106 U/mg, were neutralized with IFN-λ3 polyclonal antibodies, were susceptible to trypsin, and retained stability within defined pH and temperature ranges. Furthermore, boIFN-λ3/λ3V18M exerted antiproliferative effects on MDBK cells without cytotoxicity at 104 U/mL. Overall, boIFN-λ3 and boIFN-λ3V18M did not differ substantially in biological activity, except for reduced glycosylation of the latter. Development of boIFN-λ3 and comparative evaluation with the mutant provide theoretical insights into antiviral mechanisms of boIFN-λs and provide material for therapeutic development.

Keywords

Bovine Interferon-λ3; Pichia pastoris; Codon optimization; Glycosylation; Antiviral activity; Antiproliferative activity

Subject

Biology and Life Sciences, Animal Science, Veterinary Science and Zoology

Copyright: This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Download PDF Download Supplementary

Comments (0)

We encourage comments and feedback from a broad range of readers. See criteria for comments and our Diversity statement.

Leave a public comment
Send a private comment to the author(s)
* All users must log in before leaving a comment
Views 0
Downloads 0
Comments 0
Metrics 0
Get PDF Supplementary Files Cite Share 0
Bookmark BibSonomy Mendeley Reddit Delicious
×
Alerts
Notify me about updates to this article or when a peer-reviewed version is published.
We use cookies on our website to ensure you get the best experience.
Read more about our cookies here.