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Heterologous Expression of Toxic White Spot Syndrome Virus (WSSV) Protein in Eengineered Escherichia coli Strains

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Abstract

Aquacultural shrimps suffer economic lost due to the white spot syndrome virus (WSSV) that is the most notorious virus for its fatality and contagion, leading to a 100% death rate on infected shrimps within 7 days. However, the infection of mechanism remains a mystery and crucial problem. To elucidate the pathogenesis of WSSV, a high abundance of protein is required to identify and characterize its functions. Therefore, the optimal WSSV355 overexpression was explored in engineered Escherichia coli strains, in particular C43(DE3) as a toxic tolerance strain remedied 40% of cell growth from BL21(DE3). Meanwhile, a trace amount of WSSV355 was observed in both strains. To optimize the codon of WSSV355 using codon adaption index (CAI), an overexpression was observed with 1.32 mg/mL in C43(DE3), while the biomass was decreased by 35%. Subsequently, the co-expression with pRARE boosted the target protein up to 1.93 mg/mL. Finally, by scaling up production of WSSV355 in the fermenter with sufficient oxygen supplied, the biomass and total and soluble protein were enhanced 67.6%, 44.9%, and 7.8% compared with that in flask condition. Herein, the current approach provides efficacious solutions to produce toxic proteins via codon usage, strain selection, and processing optimization by alleviating the burden and boosting protein production in E. coli.

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References

  1. Durand, S., Lightner, D. V., Nunan, L. M., Redman, R. M., Mari, J., & Bonami, J. R. (1996). Application of gene probes as diagnostic tools for white spot baculovirus (WSBV) of penaeid shrimp. Diseases of Aquatic Organisms, 27(1), 59–66.

    Article  CAS  Google Scholar 

  2. Kumar, R., Huang, J. Y., Ng, Y. S., Chen, C. Y., & Wang, H. C. (2022). The regulation of shrimp metabolism by the white spot syndrome virus (WSSV). Reviews in Aquaculture, 14(3), 1150–1169.

    Article  Google Scholar 

  3. Li, C., Weng, S., & He, J. (2019). WSSV–host interaction: Host response and immune evasion. Fish & Shellfish Immunology, 84, 558–571.

    Article  CAS  Google Scholar 

  4. Patil, P. K., Geetha, R., Ravisankar, T., Avunje, S., Solanki, H. G., Abraham, T. J., Vinoth, S. P., Jithendran, K. P., Alavandi, S. V., & Vijayan, K. K. (2021). Economic loss due to diseases in Indian shrimp farming with special reference to Enterocytozoon hepatopenaei (EHP) and white spot syndrome virus (WSSV). Aquaculture, 533, 736231.

    Article  Google Scholar 

  5. Liu, L. K., Liu, M. J., Li, D. L., & Liu, H. P. (2021). Recent insights into anti-WSSV immunity in crayfish. Developmental & Comparative Immunology, 116, 103947.

    Article  CAS  Google Scholar 

  6. Xu, T., Shan, X., Li, Y., Yang, T., Teng, G., Wu, Q., Wang, C., Tang, K. F. J., Zhang, Q., & Jin, X. (2021). White spot syndrome virus (WSSV) prevalence in wild crustaceans in the Bohai Sea. Aquaculture, 542, 736810.

    Article  CAS  Google Scholar 

  7. Oakey, H. J., & Smith, C. S. (2018). Complete genome sequence of a white spot syndrome virus associated with a disease incursion in Australia. Aquaculture, 484, 152–159.

    Article  CAS  Google Scholar 

  8. Leu, J. H., Yang, F., Zhang, X., Xu, X., Kou, G. H., & Lo, C. F. (2009). Whispovirus. Lesser known large dsDNA. Viruses, 197–227.

  9. Xie, X., Li, H., Xu, L., & Yang, F. (2005). A simple and efficient method for purification of intact white spot syndrome virus (WSSV) viral particles. Virus Research, 108(1–2), 63–67.

    Article  CAS  PubMed  Google Scholar 

  10. Alemdar, S., Hartwig, S., Frister, T., König, J. C., Scheper, T., & Beutel, S. (2016). Heterologous expression, purification, and biochemical characterization of α-humulene synthase from Zingiber zerumbet Smith. Applied Biochemistry and Biotechnology, 178(3), 474–489.

    Article  CAS  PubMed  Google Scholar 

  11. Mahamad, P., Boonchird, C., & Panbangred, W. (2016). High level accumulation of soluble diphtheria toxin mutant (CRM197) with co-expression of chaperones in recombinant Escherichia coli. Applied Microbiology and Biotechnology, 100(14), 6319–6330.

    Article  CAS  PubMed  Google Scholar 

  12. Ajayi, B. O., Olajide, T. A., & Olayinka, E. T. (2022). 6-Gingerol attenuates pulmonary inflammation and oxidative stress in mice model of house dust mite-induced asthma. Advances in Redox Research, 5, 100036.

    Article  CAS  Google Scholar 

  13. Gumpena, R., Lountos, G. T., & Waugh, D. S. (2018). MBP-binding DARPins facilitate the crystallization of an MBP fusion protein. Acta Crystallographica Section F: Structural Biology Communications, 74(9), 549–557.

    CAS  PubMed  PubMed Central  Google Scholar 

  14. Kim, D. S., Kim, S. W., Song, J. M., Kim, S. Y., & Kwon, K. C. (2019). A new prokaryotic expression vector for the expression of antimicrobial peptide abaecin using SUMO fusion tag. BMC biotechnology, 19(1), 1–12.

    Article  PubMed  PubMed Central  Google Scholar 

  15. Hemmati, S., & Ranjbari, J. (2019). Soluble form production of recombinant human insulin-like growth factor-1 by NusA fusion partner in E. coli. Trends in Peptide and Protein Sciences, 4, 1–5.

    Google Scholar 

  16. Yu, T. H., Tan, S. I., Yi, Y. C., Xue, C., Ting, W. W., Chang, J. J., & Ng, I. S. (2022). New insight into the codon usage and medium optimization toward stable and high-level 5-aminolevulinic acid production in Escherichia coli. Biochemical Engineering Journal, 177, 108259.

    Article  CAS  Google Scholar 

  17. Fu, H., Liang, Y., Zhong, X., Pan, Z., Huang, L., Zhang, H., Xu, Y., Zhou, W., & Liu, Z. (2020). Codon optimization with deep learning to enhance protein expression. Scientific Reports, 10(1), 1–9.

    Article  Google Scholar 

  18. Wang, H. C., Wang, H. C., Kou, G. H., Lo, C. F., & Huang, W. P. (2007). Identification of icp11, the most highly expressed gene of shrimp white spot syndrome virus (WSSV). Diseases of Aquatic Organisms, 74(3), 179–189.

    Article  CAS  PubMed  Google Scholar 

  19. Ting, W. W., Yu, J. Y., Lin, Y. C., & Ng, I. S. (2022). Enhanced recombinant carbonic anhydrase in T7RNAP-equipped Escherichia coli W3110 for carbon capture storage and utilization (CCSU). Bioresource Technology, 128010.

  20. Teufel, F., Armenteros, J. J. A., Johansen, A. R., Gíslason, M. H., Pihl, S. I., Tsirigos, K. D., Winther, O., Brunak, S., Heijne, G., & Nielsen, H. (2022). SignalP 6.0 predicts all five types of signal peptides using protein language models. Nature Biotechnology, 40, 1023–1025.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Effendi, S. S. W., Tan, S. I., Ting, W. W., & Ng, I. S. (2021). Genetic design of co-expressed Mesorhizobium loti carbonic anhydrase and chaperone GroELS to enhancing carbon dioxide sequestration. International Journal of Biological Macromolecules, 167, 326–334.

    Article  CAS  PubMed  Google Scholar 

  22. Zhang, Z. X., Nong, F. T., Wang, Y. Z., Yan, C. X., Gu, Y., Song, P., & Sun, X. M. (2022). Strategies for efficient production of recombinant proteins in Escherichia coli: Alleviating the host burden and enhancing protein activity. Microbial Cell Factories, 21, 191.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Ting, W. W., Tan, S. I., & Ng, I. S. (2020). Development of chromosome-based T7 RNA polymerase and orthogonal T7 promoter circuit in Escherichia coli W3110 as a cell factory. Bioresources and Bioprocessing, 7, 54.

    Article  Google Scholar 

  24. Mittal, P., Brindle, J., Stephen, J., Plotkin, J. B., & Kudla, G. (2018). Codon usage influences fitness through RNA toxicity. Proceedings of the National Academy of Sciences, 115(34), 8639–8644.

    Article  CAS  Google Scholar 

  25. Schlegel, S., Genevaux, P., & de Gier, J. W. (2015). De-convoluting the genetic adaptations of E. coli C41 (DE3) in real time reveals how alleviating protein production stress improves yields. Cell Reports, 10(10), 1758–1766.

    Article  CAS  PubMed  Google Scholar 

  26. Effendi, S. S. W., Xue, C., Tan, S. I., & Ng, I. S. (2021). Whole-cell biocatalyst of recombinant tyrosine ammonia lyase with fusion protein and integrative chaperone in Escherichia coli for high-level p-Coumaric acid production. Journal of the Taiwan Institute of Chemical Engineers, 128, 64–72.

    Article  CAS  Google Scholar 

  27. Liu, Y. (2020). A code within the genetic code: Codon usage regulates co-translational protein folding. Cell Communication and Signaling, 18(1), 1–9.

    Article  Google Scholar 

  28. Silva, F., Queiroz, J. A., & Domingues, F. C. (2012). Evaluating metabolic stress and plasmid stability in plasmid DNA production by Escherichia coli. Biotechnology Advances, 30, 691–708.

    Article  CAS  PubMed  Google Scholar 

  29. Yi, Y. C., Xue, C., & Ng, I. S. (2021). Low-carbon-footprint production of high-end 5-aminolevulinic acid via integrative strain engineering and RuBisCo-equipped Escherichia coli. ACS Sustainable Chemistry & Engineering, 9(46), 15623–15633.

    Article  CAS  Google Scholar 

  30. Takahashi, M., & Aoyagi, H. (2020). Analysis and effect of conventional flasks in shaking culture of Escherichia coli. AMB Express, 10(1), 1–6.

    Article  Google Scholar 

  31. Crofts, A. A., Giovanetti, S. M., Rubin, E. J., Poly, F. M., Gutiérrez, R. L., Talaat, K. R., Porter, C. K., Mark, S., DeNearing, B., Brubaker, J., Maciel, M., Jr., Alcala, A. N., Chakraborty, S., Prouty, M. G., Savarino, S. J., Davies, B. W., & Trent, M. S. (2018). Enterotoxigenic E. coli virulence gene regulation in human infections. Proceedings of the National Academy of Sciences, 115(38), E8968–E8976.

    Article  CAS  Google Scholar 

  32. Akisue, R. A., Horta, A. C., & de Sousa Jr, R. (2018). Development of a fuzzy system for dissolved oxygen control in a recombinant Escherichia coli cultivation for heterologous protein expression. In Computer Aided Chemical Engineering (Vol. 43, pp. 1129–1134). Elsevier.

  33. Saito, Y., Kitagawa, W., Kumagai, T., Tajima, N., Nishimiya, Y., Tamano, K., Yasutake, Y., Tamura, T., & Kameda, T. (2019). Developing a codon optimization method for improved expression of recombinant proteins in actinobacteria. Scientific Reports, 9(1), 1–10.

    Article  Google Scholar 

  34. Mauro, V. P. (2018). Codon optimization in the production of recombinant biotherapeutics: Potential risks and considerations. BioDrugs, 32(1), 69–81.

    Article  CAS  PubMed  Google Scholar 

  35. Huang, P., Shi, J., Sun, Q., Dong, X., & Zhang, N. (2018). Engineering Pichia pastoris for efficient production of a novel bifunctional Strongylocentrotus purpuratus invertebrate-type lysozyme. Applied Biochemistry and Biotechnology, 186(2), 459–475.

    Article  CAS  PubMed  Google Scholar 

  36. Coenen, A., Marti, V. G., Müller, K., Sheremetiev, M., Finamore, L., & Schörken, U. (2022). Synthesis of polymer precursor 12-oxododecenoic acid utilizing recombinant papaya hydroperoxide lyase in an enzyme cascade. Applied Biochemistry and Biotechnology, 1–19.

  37. Yang, H., Jiang, Y., Lu, K., Xiong, H., Zhang, Y., & Wei, W. (2021). Herbicide atrazine exposure induce oxidative stress, immune dysfunction and WSSV proliferation in red swamp crayfish Procambarus clarkii. Chemosphere, 283, 131227.

    Article  CAS  PubMed  Google Scholar 

  38. Jiang, H. F., Chen, C., Jiang, X. Y., Shen, J. L., Ling, F., Li, P. F., & Wang, G. X. (2022). Luteolin in Lonicera japonica inhibits the proliferation of white spot syndrome virus in the crayfish Procambarus clarkii. Aquaculture, 550, 737852.

    Article  CAS  Google Scholar 

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Funding

The study was financially supported by the Ministry of Science and Technology (MOST 110–2221-E-006–030-MY3 and MOST 111–2221-E-006–012-MY3) in Taiwan.

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Authors and Affiliations

  1. Department of Chemical Engineering, National Cheng Kung University, Tainan, 70101, Taiwan

    Po-Yen Chen, Ying-Chen Yi & I-Son Ng

  2. Department of Biotechnology and Bioindustry Sciences, National Cheng Kung University, Tainan, 70101, Taiwan

    Han-Ching Wang

  3. International Center for the Scientific Development of Shrimp Aquaculture, National Cheng Kung University, Tainan, 70101, Taiwan

    Han-Ching Wang

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  1. Po-Yen Chen
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Contributions

Ying-Chen Yi and I-Son Ng conceived the study. Po-Yen Chen performed all the experiments and sketched the original draft. Han-Ching Wang provided the original DNA of WSSV355. I-Son Ng did methodology validation, supervised the experiments, reviewed, and edited the manuscript.

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Correspondence to I-Son Ng.

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Chen, PY., Yi, YC., Wang, HC. et al. Heterologous Expression of Toxic White Spot Syndrome Virus (WSSV) Protein in Eengineered Escherichia coli Strains. Appl Biochem Biotechnol 195, 4524–4536 (2023). https://doi.org/10.1007/s12010-023-04369-1

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