RESEARCH ARTICLE


Expression and Characterization of Rice-produced Recombinant Porcine Lactoferrin and its Antioxidant Activities



Kuan-Chih Lee1, Kun-Ting Hsieh1, Ray-Bin Chen1, Wei-Chih Lin2, Chang-Sheng Wang3, 4, Tzu-Tai Lee2, *, Liang-Jwu Chen1, 4, *
1 Institute of Molecular Biology, National Chung Hsing University, Taichung, 402 Taiwan
2 Department of Animal Science, National Chung Hsing University, Taichung, 402 Taiwan
3 Department of Agronomy, National Chung Hsing University, Taichung, 402 Taiwan
4 Agricultural Biotechnology Center, National Chung Hsing University, Taichung 402, Taiwan


Article Metrics

CrossRef Citations:
0
Total Statistics:

Full-Text HTML Views: 406
Abstract HTML Views: 126
PDF Downloads: 0
Total Views/Downloads: 532
Unique Statistics:

Full-Text HTML Views: 256
Abstract HTML Views: 83
PDF Downloads: 0
Total Views/Downloads: 339



Creative Commons License
© 2020 Lee et al.

open-access license: This is an open access article distributed under the terms of the Creative Commons Attribution 4.0 International Public License (CC-BY 4.0), a copy of which is available at: (https://creativecommons.org/licenses/by/4.0/legalcode). This license permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

* Address correspondence to this author at the Institute of Molecular Biology, National Chung Hsing University, Taichung, 402 Taiwan; Tel: +886-4-22851885; +886-4-22840366; E-mails: ljchen@nchu.edu.tw, ttlee@nchu.edu.tw


Abstract

Background:

Lactoferrin (LF) exhibits multiple beneficial biological activities and thus has been used as a health food and additive. To broaden its application in the food industry, the porcine LF (pLF) gene has been engineered into rice to produce recombinant LF (rpLF) for use as a food additive. The iron-binding and antimicrobial activities of rpLF and its positive effects on early weaned piglets have been previously evaluated, yet several features, such as the signal peptide removal, glycosylation sites and antioxidant activity of rpLF, have not been fully characterized.

Objective:

In this work, the rice-produced rpLF was purified and its biochemical structure and antioxidant activities characterized.

Methods:

HPLC, Western blot, PAS/VVL/PNA staining, Edman degradation assay, MALDI-TOF, LC-MS/MS and antioxidant activity assays were performed.

Results:

The results showed that this purified rpLF is a mature form of LF; its signal peptide was correctly removed, and two N-glycosylation sites located at N365 and N472 were identified. The molecular mass heterogeneity of rpLF could be eliminated by treatment with PNGase glycosidase, suggesting that different degrees of N-glycosylation occur in rpLF. A series of assays including the iron chelating activity, reducing power assay, lipid peroxidase activity and radical-scavenging activity showed that the antioxidant activity of rice-produced rpLF was equivalent to that of bovine LF.

Conclusion:

Rice-produced rpLF was correctly processed post-translationally and displayed antioxidant activity equivalent to that of bovine LF; thus, rice-produced rpLF can be recognized as a plant-based antioxidant to be used as a functional additive in animal feed and for the food industry.

Keywords: Recombinant porcine lactoferrin, Transgenic rice, N-glycosylation, Antioxidant activity, Western blot, Edman degradation assay, The rice-produced rpLF, Peptide.