Engineering Escherichia coli for constitutive production of monophosphoryl lipid A vaccine adjuvant.
adjuvant
adjuvant production
lipid A 1-phosphatase
lipopolysaccharide biosynthetic pathway
metabolic engineering
monophosphoryl lipid A
Journal
Biotechnology and bioengineering
ISSN: 1097-0290
Titre abrégé: Biotechnol Bioeng
Pays: United States
ID NLM: 7502021
Informations de publication
Date de publication:
07 Jan 2024
07 Jan 2024
Historique:
revised:
14
11
2023
received:
05
10
2023
accepted:
12
12
2023
medline:
7
1
2024
pubmed:
7
1
2024
entrez:
7
1
2024
Statut:
aheadofprint
Résumé
During the COVID-19 pandemic, expedient vaccine production has been slowed by the shortage of safe and effective raw materials, such as adjuvants, essential components to enhance the efficacy of vaccines. Monophosphoryl lipid A (MPLA) is a potent and safe adjuvant used in human vaccines, including the Shingles vaccine, Shingrix. 3-O-desacyl-4'-monophosphoryl lipid A (MPL), a representative MPLA adjuvant commercialized by GSK, was prepared via chemical conversion of precursors isolated from Salmonella typhimurium R595. However, the high price of these materials limits their use in premium vaccines. To combat the scarcity and high cost of safe raw materials for vaccines, we need to develop a feasible MPLA production method that is easily scaled up to meet industrial requirements. In this study, we engineered peptidoglycan and outer membrane biosynthetic pathways in Escherichia coli and developed a Escherichia coli strain, KHSC0055, that constitutively produces EcML (E. coli-produced monophosphoryl lipid A) without additives such as antibiotics or overexpression inducers. EcML production was optimized on an industrial scale via high-density fed-batch fermentation, and obtained 2.7 g of EcML (about 135,000 doses of vaccine) from a 30-L-scale fermentation. Using KHSC0055, we simplified the production process and decreased the production costs of MPLA. Then, we applied EcML purified from KHSC0055 as an adjuvant for a COVID-19 vaccine candidate (EuCorVac-19) currently in clinical trial stage III in the Philippines. By probing the efficacy and safety of EcML in humans, we established KHSC0055 as an efficient cell factory for MPLA adjuvant production.
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Subventions
Organisme : KIST intramural grants
Organisme : National Research Foundation of Korea (NRF)
ID : 2020R1A2C2100669
Organisme : Korea Health Technology R&D Project through the Korea Health Industry Development Institute (KHIDI)
ID : HV20C0007
Informations de copyright
© 2023 Wiley Periodicals LLC.
Références
Baba, T., Ara, T., Hasegawa, M., Takai, Y., Okumura, Y., Baba, M., Datsenko, K. A., Tomita, M., Wanner, B. L., & Mori, H. (2006). Construction of Escherichia coli K-12 in-frame, single-gene knockout mutants: The Keio collection. Molecular Systems Biology, 2(1), 2006.0008. https://doi.org/10.1038/msb4100050
Bentala, H., Verweij, W. R., der Vlag, A. H. V., van Loenen-Weemaes, A. M., Meijer, D. K. F., & Poelstra, K. (2002). Removal of phosphate from lipid A as a strategy to detoxify lipopolysaccharide. Shock, 18(6), 561-566. https://doi.org/10.1097/00024382-200212000-00013
Bligh, E. G., & Dyer, W. J. (1959). A rapid method of total lipid extraction and purification. Canadian Journal of Biochemistry and Physiology, 37(8), 911-917. https://www.ncbi.nlm.nih.gov/pubmed/13671378
Chung, H. S., & Raetz, C. R. H. (2010). Interchangeable domains in the Kdo transferases of Escherichia coli and Haemophilus influenzae. Biochemistry, 49(19), 4126-4137. https://doi.org/10.1021/bi100343e
Coler, R. N., Bertholet, S., Moutaftsi, M., Guderian, J. A., Windish, H. P., Baldwin, S. L., Laughlin, E. M., Duthie, M. S., Fox, C. B., Carter, D., Friede, M., Vedvick, T. S., & Reed, S. G. (2011). Development and characterization of synthetic glucopyranosyl lipid adjuvant system as a vaccine adjuvant. PLoS ONE, 6(1), e16333. https://doi.org/10.1371/journal.pone.0016333
Datsenko, K. A., & Wanner, B. L. (2000). One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products. Proceedings of the National Academy of Sciences, 97(12), 6640-6645. https://doi.org/10.1073/pnas.120163297
Garçon, N., & Di Pasquale, A. (2017). From discovery to licensure, the adjuvant system story. Human Vaccines & Immunotherapeutics, 13(1), 19-33. https://doi.org/10.1080/21645515.2016.1225635
Ghachi, M. E., Derbise, A., Bouhss, A., & Mengin-Lecreulx, D. (2005). Identification of multiple genes encoding membrane proteins with undecaprenyl pyrophosphate phosphatase (UppP) activity in Escherichia coli. Journal of Biological Chemistry, 280(19), 18689-18695. https://doi.org/10.1074/jbc.M412277200
Grant, S. G., Jessee, J., Bloom, F. R., & Hanahan, D. (1990). Differential plasmid rescue from transgenic mouse DNAs into Escherichia coli methylation-restriction mutants. Proceedings of the National Academy of Sciences 87 (12), 4645-4649. https://doi.org/10.1073/pnas.87.12.4645
Guzman, L. M., Belin, D., Carson, M. J., & Beckwith, J. (1995). Tight regulation, modulation, and high-level expression by vectors containing the arabinose PBAD promoter. Journal of Bacteriology, 177(14), 4121-4130. https://doi.org/10.1128/jb.177.14.4121-4130.1995
Hagen, S. R., Thompson, J. D., Snyder, D. S., & Myers, K. R. (1997). Analysis of a monophosphoryl lipid A immunostimulant preparation from Salmonella minnesota R595 by high-performance liquid chromatography. Journal of Chromatography A, 767(1), 53-61. https://doi.org/10.1016/S0021-9673(97)00041-1
Hankins, J. V., Madsen, J. A., Giles, D. K., Childers, B. M., Klose, K. E., Brodbelt, J. S., & Trent, M. S. (2011). Elucidation of a novel Vibrio cholerae lipid A secondary hydroxy-acyltransferase and its role in innate immune recognition. Molecular Microbiology, 81(5), 1313-1329. https://doi.org/10.1111/j.1365-2958.2011.07765.x
Ismaili, J., Rennesson, J., Aksoy, E., Vekemans, J., Vincart, B., Amraoui, Z., Van Laethem, F., Goldman, M., & Dubois, P. M. (2002). Monophosphoryl lipid A activates both human dendritic cells and T cells. The Journal of Immunology, 168(2), 926-932. https://doi.org/10.4049/jimmunol.168.2.926
Jeong, Y., Kim, G. B., Ji, Y., Kwak, G. J., Nam, G. H., Hong, Y., Kim, S., An, J., Kim, S. H., Yang, Y., Chung, H. S., & Kim, I. S. (2020). Dendritic cell activation by an E. coli-derived monophosphoryl lipid A enhances the efficacy of PD-1 blockade. Cancer Letters, 472, 19-28. https://doi.org/10.1016/j.canlet.2019.12.012
Ji, Y., An, J., Hwang, D., Ha, D. H., Lim, S. M., Lee, C., Zhao, J., Song, H. K., Yang, E. G., Zhou, P., & Chung, H. S. (2020). Metabolic engineering of Escherichia coli to produce a monophosphoryl lipid A adjuvant. Metabolic Engineering, 57, 193-202. https://doi.org/10.1016/j.ymben.2019.11.009
Johnson, A. G., Tomai, M., Solem, L., Beck, L., & Ribi, E. (1987). Characterization of a nontoxic monophosphoryl lipid A. Clinical Infectious Diseases, 9(Suppl. 5), S512-S516. https://doi.org/10.1093/clinids/9.supplement_5.s512.
Kiino, D. R., Phillips, G. J., & Silhavy, T. J. (1990). Increased expression of the bifunctional protein PrlF suppresses overproduction lethality associated with exported beta-galactosidase hybrid proteins in Escherichia coli. Journal of Bacteriology, 172(1), 185-192. https://doi.org/10.1128/jb.172.1.185-192.1990
Kramer, R. M., Archer, M. C., Orr, M. T., Dubois Cauwelaert, N., Beebe, E. A., Huang, P. D., Dowling, Q. M., Schwartz, A. M., Fedor, D. M., Vedvick, T. S., & Fox, C. B. (2018). Development of a thermostable nanoemulsion adjuvanted vaccine against tuberculosis using a design-of-experiments approach. International Journal of Nanomedicine, 13, 3689-3711. https://doi.org/10.2147/IJN.S159839
Lengeler, J. (1975). Nature and properties of hexitol transport systems in Escherichia coli. Journal of Bacteriology, 124(1), 39-47. https://doi.org/10.1128/jb.124.1.39-47.1975
Lovell, J. F., Baik, Y. O., Choi, S. K., Lee, C., Lee, J. Y., Miura, K., Huang, W. C., Park, Y. S., Woo, S. J., Seo, S. H., Kim, J. O., Song, M., Kim, C. J., Choi, J. K., Kim, J., Choo, E. J., & Choi, J. H. (2022). Interim analysis from a phase 2 randomized trial of EuCorVac-19: A recombinant protein SARS-CoV-2 RBD nanoliposome vaccine. BMC Medicine, 20(1), 462. https://doi.org/10.1186/s12916-022-02661-1
Mamat, U., Meredith, T. C., Aggarwal, P., Kühl, A., Kirchhoff, P., Lindner, B., Hanuszkiewicz, A., Sun, J., Holst, O., & Woodard, R. W. (2008). Single amino acid substitutions in either YhjD or MsbA confer viability to 3-deoxy-d-manno-oct-2-ulosonic acid-depleted Escherichia coli. Molecular Microbiology, 67(3), 633-648. https://doi.org/10.1111/j.1365-2958.2007.06074.x
Menart, V., Jevševar, S., Vilar, M., Trobiš, A., & Pavko, A. (2003). Constitutive versus thermoinducible expression of heterologous proteins in Escherichia coli based on strong PR,PL promoters from phage lambda. Biotechnology and Bioengineering, 83(2), 181-190. https://doi.org/10.1002/bit.10660
Myers, K. R., & SNYDER, D. S. (2016). Methods for the production of 3-O-deactivated-4′-monophosphoryl lipid A (3D-MLA). US Patent No. 9512159B2.
Needham, B. D., Carroll, S. M., Giles, D. K., Georgiou, G., Whiteley, M., & Trent, M. S. (2013). Modulating the innate immune response by combinatorial engineering of endotoxin. Proceedings of the National Academy of Sciences, 110(4), 1464-1469. https://doi.org/10.1073/pnas.1218080110
Orr, M. T., Fox, C. B., Baldwin, S. L., Sivananthan, S. J., Lucas, E., Lin, S., Phan, T., Moon, J. J., Vedvick, T. S., Reed, S. G., & Coler, R. N. (2013). Adjuvant formulation structure and composition are critical for the development of an effective vaccine against tuberculosis. Journal of Controlled Release, 172(1), 190-200. https://doi.org/10.1016/j.jconrel.2013.07.030
Qureshi, N., Takayama, K., & Ribi, E. (1982). Purification and structural determination of nontoxic lipid A obtained from the lipopolysaccharide of Salmonella typhimurium. Journal of Biological Chemistry, 257(19), 11808-11815.
Rong Fu Wang, W., & Kushner, S. R. (1991). Construction of versatile low-copy-number vectors for cloning, sequencing and gene expression in Escherichia coli. Gene, 100, 195-199. https://doi.org/10.1016/0378-1119(91)90366-J
Sanina, N. (2019). Vaccine adjuvants derived from marine organisms. Biomolecules, 9(8), 340. https://doi.org/10.3390/biom9080340
Snyder, W. B., & Silhavy, T. J. (1992). Enhanced export of beta-galactosidase fusion proteins in prlF mutants is lon dependent. Journal of Bacteriology, 174(17), 5661-5668. https://doi.org/10.1128/jb.174.17.5661-5668.1992
Steimle, A., Autenrieth, I. B., & Frick, J. S. (2016). Structure and function: Lipid A modifications in commensals and pathogens. International Journal of Medical Microbiology, 306(5), 290-301. https://doi.org/10.1016/j.ijmm.2016.03.001
Takayama, K., Qureshi, N., Ribi, E., & Cantrell, J. L. (1984). Separation and characterization of toxic and nontoxic forms of lipid A. Clinical Infectious Diseases, 6(4), 439-443. https://doi.org/10.1093/clinids/6.4.439
Thomason, L. C., Costantino, N., & Court, D. L. (2007). E. coli genome manipulation by P1 transduction. Current Protocols in Molecular Biology, 79(1), 1.17.11-11.17.18. https://doi.org/10.1002/0471142727.mb0117s79
Wang, B., Han, Y., Li, Y., Li, Y., & Wang, X. (2015). Immuno-stimulatory activity of Escherichia coli mutants producing Kdo2-Monophosphoryl-Lipid A or Kdo2-Pentaacyl-Monophosphoryl-Lipid A. PLoS ONE, 10(12), e0144714. https://doi.org/10.1371/journal.pone.0144714
Wang, Y. Q., Bazin-Lee, H., Evans, J. T., Casella, C. R., & Mitchell, T. C. (2020). MPL adjuvant contains competitive antagonists of human TLR4. Frontiers in Immunology, 11, 577823. https://doi.org/10.3389/fimmu.2020.577823
Wang, Z., Zhao, A., Wang, C., Huang, D., Yu, J., Yu, L., Wu, Y., & Wang, X. (2023). Metabolic engineering of Escherichia coli to efficiently produce monophosphoryl lipid A. Biotechnology and Applied Biochemistry, 70(3), 1332-1345. https://doi.org/10.1002/bab.2443
Yu, D., Ellis, H. M., Lee, E.-C., Jenkins, N. A., Copeland, N. G., & Court, D. L. (2000). An efficient recombination system for chromosome engineering in Escherichia coli. Proceedings of the National Academy of Sciences, 97(11), 5978-5983. https://doi.org/10.1073/pnas.100127597
Zhao, J., An, J., Hwang, D., Wu, Q., Wang, S., Gillespie, R. A., Yang, E. G., Guan, Z., Zhou, P., & Chung, H. S. (2019). The lipid A 1-Phosphatase, LpxE, functionally connects multiple layers of bacterial envelope biogenesis. mBio, 10(3), e00886-19. https://doi.org/10.1128/mBio.00886-19