High-Level Secretory Production of Recombinant E2-Spy Antigen Protein via Combined Strategy in <i>Pichia pastoris</i>
E2-Spy (abbreviated as ES) plays a vital role as a component in the Bacterial-Like Particles (BLPs) vaccine against classical swine fever virus (CSFV). This vaccine demonstrates remarkable immunoprotection, highlighting the importance of augmenting ES production in the development of CSFV subunit va...
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| Format: | Article |
| Language: | English |
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MDPI AG
2024-02-01
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| Series: | Fermentation |
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| Online Access: | https://www.mdpi.com/2311-5637/10/2/99 |
| _version_ | 1797298283647860736 |
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| author | Bingkun Li Yiheng Zheng Shida Zhao Yaohan Zhang Ding Li |
| author_facet | Bingkun Li Yiheng Zheng Shida Zhao Yaohan Zhang Ding Li |
| author_sort | Bingkun Li |
| collection | DOAJ |
| description | E2-Spy (abbreviated as ES) plays a vital role as a component in the Bacterial-Like Particles (BLPs) vaccine against classical swine fever virus (CSFV). This vaccine demonstrates remarkable immunoprotection, highlighting the importance of augmenting ES production in the development of CSFV subunit vaccines. In this study, a <i>Pichia pastoris</i> strain capable of high-yield secretory production of ES was developed through signal peptide engineering, gene dosage optimization and co-expression of molecular chaperones. Initially, a hybrid signal peptide cSP3 was engineered, leading to a 3.38-fold increase in ES production when compared to the control strain 1-α-ES. Subsequently, cSP3 was evaluated for its expression efficiency alongside different commonly used signal peptides under multicopy conditions. SDS-PAGE analysis revealed that 2-αd14-ES exhibited the highest ES production, displaying a 4.38-fold increase in comparison to 1-α-ES. Afterwards, SSA1, YDJ1, BIP, LHS1, and their combinations were integrated into 2-αd14-ES, resulting in a 1.92-fold rise in ES production compared to 2-αd14-ES (equivalent to a 6.18-fold increase compared to 1-α-ES). The final yield of ES was evaluated as 168.3 mg/L through comparison with serially diluted BSA protein bands. |
| first_indexed | 2024-03-07T22:33:40Z |
| format | Article |
| id | doaj.art-7e7188f5e8ef4485b67007e016e97dce |
| institution | Directory Open Access Journal |
| issn | 2311-5637 |
| language | English |
| last_indexed | 2024-03-07T22:33:40Z |
| publishDate | 2024-02-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Fermentation |
| spelling | doaj.art-7e7188f5e8ef4485b67007e016e97dce2024-02-23T15:15:56ZengMDPI AGFermentation2311-56372024-02-011029910.3390/fermentation10020099High-Level Secretory Production of Recombinant E2-Spy Antigen Protein via Combined Strategy in <i>Pichia pastoris</i>Bingkun Li0Yiheng Zheng1Shida Zhao2Yaohan Zhang3Ding Li4Institute of Veterinary Immunology & Engineering, Jiangsu Academy of Agricultural Sciences, Nanjing 210014, ChinaInstitute of Veterinary Immunology & Engineering, Jiangsu Academy of Agricultural Sciences, Nanjing 210014, ChinaInstitute of Veterinary Immunology & Engineering, Jiangsu Academy of Agricultural Sciences, Nanjing 210014, ChinaInstitute of Veterinary Immunology & Engineering, Jiangsu Academy of Agricultural Sciences, Nanjing 210014, ChinaInstitute of Veterinary Immunology & Engineering, Jiangsu Academy of Agricultural Sciences, Nanjing 210014, ChinaE2-Spy (abbreviated as ES) plays a vital role as a component in the Bacterial-Like Particles (BLPs) vaccine against classical swine fever virus (CSFV). This vaccine demonstrates remarkable immunoprotection, highlighting the importance of augmenting ES production in the development of CSFV subunit vaccines. In this study, a <i>Pichia pastoris</i> strain capable of high-yield secretory production of ES was developed through signal peptide engineering, gene dosage optimization and co-expression of molecular chaperones. Initially, a hybrid signal peptide cSP3 was engineered, leading to a 3.38-fold increase in ES production when compared to the control strain 1-α-ES. Subsequently, cSP3 was evaluated for its expression efficiency alongside different commonly used signal peptides under multicopy conditions. SDS-PAGE analysis revealed that 2-αd14-ES exhibited the highest ES production, displaying a 4.38-fold increase in comparison to 1-α-ES. Afterwards, SSA1, YDJ1, BIP, LHS1, and their combinations were integrated into 2-αd14-ES, resulting in a 1.92-fold rise in ES production compared to 2-αd14-ES (equivalent to a 6.18-fold increase compared to 1-α-ES). The final yield of ES was evaluated as 168.3 mg/L through comparison with serially diluted BSA protein bands.https://www.mdpi.com/2311-5637/10/2/99E2-Spyhybrid signal peptide cSP3gene dosage optimizationmolecular chaperone |
| spellingShingle | Bingkun Li Yiheng Zheng Shida Zhao Yaohan Zhang Ding Li High-Level Secretory Production of Recombinant E2-Spy Antigen Protein via Combined Strategy in <i>Pichia pastoris</i> Fermentation E2-Spy hybrid signal peptide cSP3 gene dosage optimization molecular chaperone |
| title | High-Level Secretory Production of Recombinant E2-Spy Antigen Protein via Combined Strategy in <i>Pichia pastoris</i> |
| title_full | High-Level Secretory Production of Recombinant E2-Spy Antigen Protein via Combined Strategy in <i>Pichia pastoris</i> |
| title_fullStr | High-Level Secretory Production of Recombinant E2-Spy Antigen Protein via Combined Strategy in <i>Pichia pastoris</i> |
| title_full_unstemmed | High-Level Secretory Production of Recombinant E2-Spy Antigen Protein via Combined Strategy in <i>Pichia pastoris</i> |
| title_short | High-Level Secretory Production of Recombinant E2-Spy Antigen Protein via Combined Strategy in <i>Pichia pastoris</i> |
| title_sort | high level secretory production of recombinant e2 spy antigen protein via combined strategy in i pichia pastoris i |
| topic | E2-Spy hybrid signal peptide cSP3 gene dosage optimization molecular chaperone |
| url | https://www.mdpi.com/2311-5637/10/2/99 |
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