文献类型：期刊文献

英文题名：Investigation of Kluyveromyces marxianus as a novel host for large-scale production of porcine parvovirus virus-like particles

作者：Yang, Deqiang[1,2];Chen, Lei[1,2];Duan, Jinkun[1,2];Yu, Yao[1,2];Zhou, Jungang[1,2];Lu, Hong[1,2,3]

机构：[1]Fudan Univ, Sch Life Sci, State Key Lab Genet Engn, 2005 Songhu Rd, Shanghai 200438, Peoples R China;[2]Shanghai Engn Res Ctr Ind Microorganisms, 2005 Songhu Rd, Shanghai 200438, Peoples R China;[3]East China Univ Sci & Technol, Shanghai Collaborat Innovat Ctr Biomfg SCICB, 130 Meilong Rd, Shanghai 200237, Peoples R China

年份：2021

卷号：20

期号：1

外文期刊名：MICROBIAL CELL FACTORIES

收录：;WOS:【SCI-EXPANDED(收录号:WOS:000611535200001)】；

基金：This study was supported by the projects of National Natural Science Foundation of China (No. 31970068 and No. 31770094 to HL), and Science and Technology Research Program of Shanghai (No. 18391901800 and No. 19DZ2282100 to HL).

语种：英文

外文关键词：Kluyveromyces marxianus; Porcine parvovirus; Virus-like particles; Vaccine; Immunity

摘要：Background: Porcine Parvovirus (PPV) is a Parvovirinae virus that can cause embryonic and fetal loss and death and mummification in affected fetal pigs. Unlike conventional vaccines, virus-like particles (VLPs) inherit the natural structure of their authentic virions and highly immunostimulatory that can induce strong humoral immune and T cell responses with no risk of pathogenicity. The production of PPV VLPs is still a challenge based on traditional expression platforms due to their low yields and high culture costs. Kluyveromyces marxianus is a safe and fast-growing eukaryote that can get high biomass with low-cost cultures. In this study, we investigated the expression and downstream processes of PPV VLPs in K. marxianus, and the potential for effective stand-alone vaccines. Results: After optimization according to the codon bias of K. marxianus, the VP2 protein from Kresse strain was highly expressed. In a 5 L fermentator, the yield of PPV VLPs reached 2.5 g/L, quantified by HPLC, using a defined mineral medium after 48 h fermentation. Two strategies were established to purify intracellular PPV VLPs: (i) Using the cation exchange chromatography coupled with Sephacryl (R) S-500 HR chromatography to purify VLPs from the supernatants of pH adjusted cell lysates. (ii) Using anion exchange chromatography followed by cross-flow diafiltration to recover the VLPs precipitated in pH adjusted cell lysates. The purity of PPV VLPs reached about 95%, and total recovery was more than 60%. Vaccination of mice with the purified PPV VLPs induced high titers of specific IgG antibodies in sera, and showed hemagglutination inhibitions on both swine and guinea pig erythrocytes. Spleen lymphocyte proliferation and cytokines detection suggested the PPV VLPs produced by K. marxianus provoked the cellular immune and humoral immunity responses in mice. Conclusions: This is the highest production of recombinant PPV VLPs achieved to date. The superiorities, Generally Recognized As Safe (GRAS), high production, short lead time, and low cost, make K. marxianus a greatly competitive platform for bioproduction of PPV VLPs vaccine.