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Advances in Synthetic Biology Tools and Engineering of Corynebacterium glutamicum as a Platform Host for Recombinant Protein Production

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Abstract

Corynebacterium glutamicum is a gram-positive and non-pathogenic microorganism that has traditionally been used as a major host for amino acid production. With the rapid increase in demand for high-value bioindustrial and pharmaceutically-relevant proteins, C. glutamicum has also gained significant interest as an attractive host for the production of recombinant proteins owing to several advantages over other microbial hosts, including Escherichia coli. As an emerging technology, various synthetic biology tools have been developed and successfully used to design and engineer C. glutamicum cells, making this microorganism a promising cell factory in the bioindustry field. In this review, we first summarize the synthetic biology tools for genome editing, synthetic parts for gene expression and regulation (i.e., promoters, ribosome binding sites, and bicistronic system), plasmids, and chassis, which have been intensively used for designing optimal expression constructs for C. glutamicum. Subsequently, as applications of synthetic biology tools, we discuss the extensive engineering of C. glutamicum as a potential platform for the secretory production of recombinant proteins.

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References

  1. Rosano, G. L. and E. A. Ceccarelli (2014) Recombinant protein expression in Escherichia coli: advances and challenges. Front. Microbiol. 5: 172.

    PubMed  PubMed Central  Google Scholar 

  2. Qian, Z. G., X. X. Xia, J. H. Choi, and S. Y. Lee (2008) Proteome-based identification of fusion partner for high-level extracellular production of recombinant proteins in Escherichia coli. Biotechnol. Bioeng. 101: 587–601.

    PubMed  CAS  Google Scholar 

  3. Dassler, T., G. Wich, and G. Schmid (2010) Process for the production of proteins by fermentation. European Patent EP1903105B1.

  4. Burdette, L. A., S. A. Leach, H. T. Wong, and D. Tullman-Ercek (2018) Developing Gram-negative bacteria for the secretion of heterologous proteins. Microb. Cell Fact. 17: 196.

    PubMed  CAS  PubMed Central  Google Scholar 

  5. Taguchi, S., T. Ooi, K. Mizuno, and H. Matsusaki (2015) Advances and needs for endotoxin-free production strains. Appl. Microbiol. Biotechnol. 99: 9349–9360.

    PubMed  CAS  Google Scholar 

  6. Freudl, R. (2017) Beyond amino acids: use of the Corynebacterium glutamicum cell factory for the secretion of heterologous proteins. J. Biotechnol. 258: 101–109.

    PubMed  CAS  Google Scholar 

  7. Kirchner, O. and A. Tauch (2003) Tools for genetic engineering in the amino acid-producing bacterium Corynebacterium glutamicum. J. Biotechnol. 104: 287–299.

    PubMed  CAS  Google Scholar 

  8. Zhang, G., X. Ren, X. Liang, Y. Wang, D. Feng, Y. Zhang, M. Xian, and H. Zou (2021) Improving the microbial production of amino acids: from conventional approaches to recent trends. Biotechnol. Bioprocess Eng. 26: 708–727.

    CAS  Google Scholar 

  9. Liu, X.-X., Y. Li, and Z.-H. Bai (2021) Corynebacterium glutamicum as a robust microbial factory for production of value-added proteins and small molecules: fundamentals and applications. pp. 235–263. In: V. Singh (ed.). Microbial Cell Factories Engineering for Production of Biomolecules. Academic Press, London, UK.

    Google Scholar 

  10. Lee, M. J. and P. Kim (2018) Recombinant protein expression system in Corynebacterium glutamicum and its application. Front. Microbiol. 9: 2523.

    PubMed  PubMed Central  Google Scholar 

  11. Liu, X., W. Zhang, Z. Zhao, X. Dai, Y. Yang, and Z. Bai (2017) Protein secretion in Corynebacterium glutamicum. Crit. Rev. Biotechnol. 37: 541–551.

    PubMed  CAS  Google Scholar 

  12. Wendisch, V. F., J. M. P. Jorge, F. Pérez-García, and E. Sgobba (2016) Updates on industrial production of amino acids using Corynebacterium glutamicum. World J. Microbiol. Biotechnol. 32: 105.

    PubMed  Google Scholar 

  13. Sauer, M. and D. Mattanovich (2012) Construction of microbial cell factories for industrial bioprocesses. J. Chem. Technol. Biotechnol. 87: 445–450.

    CAS  Google Scholar 

  14. Sasikumar, K., S. Hannibal, V. F. Wendisch, and K. M. Nampoothiri (2021) Production of biopolyamide precursors 5-amino valeric acid and putrescine from rice straw hydrolysate by engineered Corynebacterium glutamicum. Front. Bioeng. Biotechnol. 9: 635509.

    PubMed  PubMed Central  Google Scholar 

  15. Cho, J. S., K. R. Choi, C. P. S. Prabowo, J. H. Shin, D. Yang, J. Jang, and S. Y. Lee (2017) CRISPR/Cas9-coupled recombineering for metabolic engineering of Corynebacterium glutamicum. Metab. Eng. 42: 157–167.

    PubMed  CAS  Google Scholar 

  16. Deng, C., X. Lv, Y. Liu, J. Li, W. Lu, G. Du, and L. Liu (2019) Metabolic engineering of Corynebacterium glutamicum S9114 based on whole-genome sequencing for efficient N-acetylglucosamine synthesis. Synth. Syst. Biotechnol. 4: 120–129. (Erratum published 2020, Synth. Syst. Biotechnol. 5: 330–331)

    PubMed  PubMed Central  Google Scholar 

  17. Xu, J., M. Han, J. Zhang, Y. Guo, and W. Zhang (2014) Metabolic engineering Corynebacterium glutamicum for the L-lysine production by increasing the flux into L-lysine biosynthetic pathway. Amino Acids 46: 2165–2175.

    PubMed  CAS  Google Scholar 

  18. Ko, Y. J., Y.-C. Joo, J. E. Hyeon, E. Lee, M.-E. Lee, J. Seok, S. W. Kim, C. Park, and S. O. Han (2018) Biosynthesis of organic photosensitizer Zn-porphyrin by diphtheria toxin repressor (DtxR)-mediated global upregulation of engineered heme biosynthesis pathway in Corynebacterium glutamicum. Sci. Rep. 8: 14460.

    PubMed  PubMed Central  Google Scholar 

  19. Kogure, T., M. Suda, K. Hiraga, and M. Inui (2021) Protocatechuate overproduction by Corynebacterium glutamicum via simultaneous engineering of native and heterologous biosynthetic pathways. Metab. Eng. 65: 232–242.

    PubMed  CAS  Google Scholar 

  20. Cheng, F., H. Yu, and G. Stephanopoulos (2019) Engineering Corynebacterium glutamicum for high-titer biosynthesis of hyaluronic acid. Metab. Eng. 55: 276–289.

    PubMed  CAS  Google Scholar 

  21. Shin, J. H., S. H. Park, Y. H. Oh, J. W. Choi, M. H. Lee, J. S. Cho, K. J. Jeong, J. C. Joo, J. Yu, S. J. Park, and S. Y. Lee (2016) Metabolic engineering of Corynebacterium glutamicum for enhanced production of 5-aminovaleric acid. Microb. Cell Fact. 15: 174.

    PubMed  PubMed Central  Google Scholar 

  22. Schäfer, A., A. Tauch, W. Jäger, J. Kalinowski, G. Thierbach, and A. Pühler (1994) Small mobilizable multi-purpose cloning vectors derived from the Escherichia coli plasmids pK18 and pK19: selection of defined deletions in the chromosome of Corynebacterium glutamicum. Gene 145: 69–73.

    PubMed  Google Scholar 

  23. Nesvera, J. and M. Pátek (2011) Tools for genetic manipulations in Corynebacterium glutamicum and their applications. Appl. Microbiol. Biotechnol. 90: 1641–1654.

    PubMed  CAS  Google Scholar 

  24. Wang, Q., J. Zhang, N. H. Al Makishah, X. Sun, Z. Wen, Y. Jiang, and S. Yang (2021) Advances and perspectives for genome editing tools of Corynebacterium glutamicum. Front. Microbiol. 12: 654058.

    PubMed  PubMed Central  Google Scholar 

  25. Tan, Y., D. Xu, Y. Li, and X. Wang (2012) Construction of a novel sacB-based system for marker-free gene deletion in Corynebacterium glutamicum. Plasmid 67: 44–52.

    PubMed  CAS  Google Scholar 

  26. Wang, T., Y. Li, J. Li, D. Zhang, N. Cai, G. Zhao, H. Ma, C. Shang, Q. Ma, Q. Xu, and N. Chen (2019) An update of the suicide plasmid-mediated genome editing system in Corynebacterium glutamicum. Microb. Biotechnol. 12: 907–919.

    PubMed  PubMed Central  Google Scholar 

  27. Sawitzke, J. A., N. Costantino, X. T. Li, L. C. Thomason, M. Bubunenko, C. Court, and D. L. Court (2011) Probing cellular processes with oligo-mediated recombination and using the knowledge gained to optimize recombineering. J. Mol. Biol. 407: 45–59.

    PubMed  CAS  PubMed Central  Google Scholar 

  28. Binder, S., S. Siedler, J. Marienhagen, M. Bott, and L. Eggeling (2013) Recombineering in Corynebacterium glutamicum combined with optical nanosensors: a general strategy for fast producer strain generation. Nucleic Acids Res. 41: 6360–6369.

    PubMed  CAS  PubMed Central  Google Scholar 

  29. Krylov, A. A., E. E. Kolontaevsky, and S. V. Mashko (2014) Oligonucleotide recombination in corynebacteria without the expression of exogenous recombinases. J. Microbiol. Methods 105: 109–115.

    PubMed  CAS  Google Scholar 

  30. Wu, M., Y. Xu, J. Yang, and G. Shang (2020) Homing endonuclease I-SceI-mediated Corynebacterium glutamicum ATCC 13032 genome engineering. Appl. Microbiol. Biotechnol. 104: 3597–3609.

    PubMed  CAS  Google Scholar 

  31. Huang, Y., L. Li, S. Xie, N. Zhao, S. Han, Y. Lin, and S. Zheng (2017) Recombineering using RecET in Corynebacterium glutamicum ATCC14067 via a self-excisable cassette. Sci. Rep. 7: 7916.

    PubMed  PubMed Central  Google Scholar 

  32. Luo, G., N. Zhao, S. Jiang, and S. Zheng (2021) Application of RecET-Cre/loxP system in Corynebacterium glutamicum ATCC14067 for L-leucine production. Biotechnol. Lett. 43: 297–306.

    PubMed  CAS  Google Scholar 

  33. Gorshkova, N. V., J. S. Lobanova, I. L. Tokmakova, S. V. Smirnov, V. Z. Akhverdyan, A. A. Krylov, and S. V. Mashko (2018) Mu-driven transposition of recombinant mini-Mu unit DNA in the Corynebacterium glutamicum chromosome. Appl. Microbiol. Biotechnol. 102: 2867–2884.

    PubMed  CAS  PubMed Central  Google Scholar 

  34. Inui, M., Y. Tsuge, N. Suzuki, A. A. Vertès, and H. Yukawa (2005) Isolation and characterization of a native composite transposon, Tn14751, carrying 17.4 kilobases of Corynebacterium glutamicum chromosomal DNA. Appl. Environ. Microbiol. 71: 407–416.

    PubMed  CAS  PubMed Central  Google Scholar 

  35. Suzuki, N., N. Okai, H. Nonaka, Y. Tsuge, M. Inui, and H. Yukawa (2006) High-throughput transposon mutagenesis of Corynebacterium glutamicum and construction of a single-gene disruptant mutant library. Appl. Environ. Microbiol. 72: 3750–3755.

    PubMed  CAS  PubMed Central  Google Scholar 

  36. Tsuge, Y., N. Suzuki, M. Inui, and H. Yukawa (2007) Random segment deletion based on IS31831 and Cre/loxP excision system in Corynebacterium glutamicum. Appl. Microbiol. Biotechnol. 74: 1333–1341.

    PubMed  CAS  Google Scholar 

  37. Jinek, M., K. Chylinski, I. Fonfara, M. Hauer, J. A. Doudna, and E. Charpentier (2012) A programmable dual-RNA-guided DNA endonuclease in adaptive bacterial immunity. Science 337: 816–821.

    PubMed  CAS  PubMed Central  Google Scholar 

  38. Pátek, M., J. Nesvera, A. Guyonvarch, O. Reyes, and G. Leblon (2003) Promoters of Corynebacterium glutamicum. J. Biotechnol. 104: 311–323.

    PubMed  Google Scholar 

  39. Jiang, Y., F. Qian, J. Yang, Y. Liu, F. Dong, C. Xu, B. Sun, B. Chen, X. Xu, Y. Li, R. Wang, and S. Yang (2017) CRISPR-Cpf1 assisted genome editing of Corynebacterium glutamicum. Nat. Commun. 8: 15179.

    PubMed  PubMed Central  Google Scholar 

  40. Liu, J., Y. Wang, Y. Lu, P. Zheng, J. Sun, and Y. Ma (2017) Development of a CRISPR/Cas9 genome editing toolbox for Corynebacterium glutamicum. Microb. Cell Fact. 16: 205.

    PubMed  PubMed Central  Google Scholar 

  41. Zhao, N., L. Li, G. Luo, S. Xie, Y. Lin, S. Han, Y. Huang, and S. Zheng (2020) Multiplex gene editing and large DNA fragment deletion by the CRISPR/Cpf1-RecE/T system in Corynebacterium glutamicum. J. Ind. Microbiol. Biotechnol. 47: 599–608.

    PubMed  CAS  Google Scholar 

  42. Liu, W., D. Tang, H. Wang, J. Lian, L. Huang, and Z. Xu (2019) Combined genome editing and transcriptional repression for metabolic pathway engineering in Corynebacterium glutamicum using a catalytically active Cas12a. Appl. Microbiol. Biotechnol. 103: 8911–8922.

    PubMed  CAS  Google Scholar 

  43. Billman-Jacobe, H., A. L. Hodgson, M. Lightowlers, P. R. Wood, and A. J. Radford (1994) Expression of ovine gamma interferon in Escherichia coli and Corynebacterium glutamicum. Appl. Environ. Microbiol. 60: 1641–1645.

    PubMed  CAS  PubMed Central  Google Scholar 

  44. Brabetz, W., W. Liebl, and K. H. Schleifer (1991) Studies on the utilization of lactose by Corynebacterium glutamicum, bearing the lactose operon of Escherichia coli. Arch. Microbiol. 155: 607–612.

    PubMed  CAS  Google Scholar 

  45. Ben-Samoun, K., G. Leblon, and O. Reyes (1999) Positively regulated expression of the Escherichia coli araBAD promoter in Corynebacterium glutamicum. FEMS Microbiol. Lett. 174: 125–130.

    PubMed  CAS  Google Scholar 

  46. Kortmann, M., V. Kuhl, S. Klaffl, and M. Bott (2015) A chromosomally encoded T7 RNA polymerase-dependent gene expression system for Corynebacterium glutamicum: construction and comparative evaluation at the single-cell level. Microb. Biotechnol. 8: 253–265.

    PubMed  CAS  Google Scholar 

  47. Liu, X., Y. Yang, W. Zhang, Y. Sun, F. Peng, L. Jeffrey, L. Harvey, B. McNeil, and Z. Bai (2016) Expression of recombinant protein using Corynebacterium Glutamicum: progress, challenges and applications. Crit. Rev. Biotechnol. 36: 652–664.

    PubMed  Google Scholar 

  48. Lu, N., C. Zhang, W. Zhang, H. Xu, Y. Li, M. Wei, J. Meng, Y. Meng, J. Wang, and N. Chen (2021) A myo-inositol-inducible expression system for Corynebacterium glutamicum and its application. Front. Bioeng. Biotechnol. 9: 746322.

    PubMed  PubMed Central  Google Scholar 

  49. Kim, M. J., S. S. Yim, J. W. Choi, and K. J. Jeong (2016) Development of a potential stationary-phase specific gene expression system by engineering of SigB-dependent cg3141 promoter in Corynebacterium glutamicum. Appl. Microbiol. Biotechnol. 100: 4473–4483.

    PubMed  CAS  Google Scholar 

  50. Huang, J., J. Chen, Y. Wang, T. Shi, X. Ni, W. Pu, J. Liu, Y. Zhou, N. Cai, S. Han, P. Zheng, and J. Sun (2021) Development of a hyperosmotic stress inducible gene expression system by engineering the MtrA/MtrB-dependent NCgl1418 promoter in Corynebacterium glutamicum. Front. Microbiol. 12: 718511.

    PubMed  PubMed Central  Google Scholar 

  51. Peyret, J. L., N. Bayan, G. Joliff, T. Gulik-Krzywicki, L. Mathieu, E. Shechter, and G. Leblon (1993) Characterization of the cspB gene encoding PS2, an ordered surface-layer protein in Corynebacterium glutamicum. Mol. Microbiol. 9: 97–109.

    PubMed  CAS  Google Scholar 

  52. An, S. J., S. S. Yim, and K. J. Jeong (2013) Development of a secretion system for the production of heterologous proteins in Corynebacterium glutamicum using the Porin B signal peptide. Protein Expr. Purif. 89: 251–257.

    PubMed  CAS  Google Scholar 

  53. Zhang, S., D. Liu, Z. Mao, Y. Mao, H. Ma, T. Chen, X. Zhao, and Z. Wang (2018) Model-based reconstruction of synthetic promoter library in Corynebacterium glutamicum. Biotechnol. Lett. 40: 819–827.

    PubMed  CAS  Google Scholar 

  54. Wei, H., Y. Ma, Q. Chen, Y. Cui, L. Du, Q. Ma, Y. Li, X. Xie, and N. Chen (2018) Identification and application of a novel strong constitutive promoter in Corynebacterium glutamicum. Ann. Microbiol. 68: 375–382.

    CAS  Google Scholar 

  55. Chai, M., C. Deng, Q. Chen, W. Lu, Y. Liu, J. Li, G. Du, X. Lv, and L. Liu (2021) Synthetic biology toolkits and metabolic engineering applied in Corynebacterium glutamicum for biomanufacturing. ACS Synth. Biol. 10: 3237–3250.

    PubMed  CAS  Google Scholar 

  56. Yim, S. S., S. J. An, M. Kang, J. Lee, and K. J. Jeong (2013) Isolation of fully synthetic promoters for high-level gene expression in Corynebacterium glutamicum. Biotechnol. Bioeng. 110: 2959–2969.

    PubMed  CAS  Google Scholar 

  57. Rytter, J. V., S. Helmark, J. Chen, M. J. Lezyk, C. Solem, and P. R. Jensen (2014) Synthetic promoter libraries for Corynebacterium glutamicum. Appl. Microbiol. Biotechnol. 98: 2617–2623.

    PubMed  CAS  Google Scholar 

  58. Yim, S. S., J. W. Choi, S. H. Lee, and K. J. Jeong (2016) Modular optimization of a hemicellulose-utilizing pathway in Corynebacterium glutamicum for consolidated bioprocessing of hemicellulosic biomass. ACS Synth. Biol. 5: 334–343.

    PubMed  CAS  Google Scholar 

  59. Zhang, B., N. Zhou, Y. M. Liu, C. Liu, C. B. Lou, C. Y. Jiang, and S. J. Liu (2015) Ribosome binding site libraries and pathway modules for shikimic acid synthesis with Corynebacterium glutamicum. Microb. Cell Fact. 14: 71.

    PubMed  PubMed Central  Google Scholar 

  60. Shi, F., M. Luan, and Y. Li (2018) Ribosomal binding site sequences and promoters for expressing glutamate decarboxylase and producing γ-aminobutyrate in Corynebacterium glutamicum. AMB Express 8: 61.

    PubMed  PubMed Central  Google Scholar 

  61. Li, N., W. Zeng, S. Xu, and J. Zhou (2020) Obtaining a series of native gradient promoter-5′-UTR sequences in Corynebacterium glutamicum ATCC 13032. Microb. Cell Fact. 19: 120.

    PubMed  PubMed Central  Google Scholar 

  62. Mutalik, V. K., J. C. Guimaraes, G. Cambray, C. Lam, M. J. Christoffersen, Q.-A. Mai, A. B. Tran, M. Paull, J. D. Keasling, A. P. Arkin, and D. Endy (2013) Precise and reliable gene expression via standard transcription and translation initiation elements. Nat. Methods 10: 354–360.

    PubMed  CAS  Google Scholar 

  63. Gießelmann, G., D. Dietrich, L. Jungmann, M. Kohlstedt, E. J. Jeon, S. S. Yim, F. Sommer, D. Zimmer, T. Mühlhaus, M. Schroda, K. J. Jeong, J. Becker, and C. Wittmann (2019) Metabolic engineering of Corynebacterium glutamicum for high-level ectoine production: design, combinatorial assembly, and implementation of a transcriptionally balanced heterologous ectoine pathway. Biotechnol. J. 14: e1800417.

    PubMed  Google Scholar 

  64. Duan, Y., W. Zhai, W. Liu, X. Zhang, J.-S. Shi, X. Zhang, and Z. Xu (2021) Fine-tuning multi-gene clusters via well-characterized gene expression regulatory elements: case study of the arginine synthesis pathway in C. glutamicum. ACS Synth. Biol. 10: 38–48.

    PubMed  Google Scholar 

  65. Sun, M., X. Gao, Z. Zhao, A. Li, Y. Wang, Y. Yang, X. Liu, and Z. Bai (2020) Enhanced production of recombinant proteins in Corynebacterium glutamicum by constructing a bicistronic gene expression system. Microb. Cell Fact. 19: 113.

    PubMed  CAS  PubMed Central  Google Scholar 

  66. Zhang, W., Z. Zhao, Y. Yang, X. Liu, and Z. Bai (2017) Construction of an expression vector that uses the aph promoter for protein expression in Corynebacterium glutamicum. Plasmid 94: 1–6.

    PubMed  CAS  Google Scholar 

  67. Eikmanns, B. J., E. Kleinertz, W. Liebl, and H. Sahm (1991) A family of Corynebacterium glutamicum/Escherichia coli shuttle vectors for cloning, controlled gene expression, and promoter probing. Gene 102: 93–98.

    PubMed  CAS  Google Scholar 

  68. Suzuki, N., K. Watanabe, N. Okibe, Y. Tsuchida, M. Inui, and H. Yukawa (2009) Identification of new secreted proteins and secretion of heterologous amylase by C. glutamicum. Appl. Microbiol. Biotechnol. 82: 491–500.

    PubMed  CAS  Google Scholar 

  69. Bakkes, P. J., P. Ramp, A. Bida, D. Dohmen-Olma, M. Bott, and R. Freudl (2020) Improved pEKEx2-derived expression vectors for tightly controlled production of recombinant proteins in Corynebacterium glutamicum. Plasmid 112: 102540.

    PubMed  Google Scholar 

  70. Li, Y., Y. Ai, J. Zhang, J. Fei, B. Liu, J. Wang, M. Li, Q. Zhao, and J. Song (2020) A novel expression vector for Corynebacterium glutamicum with an auxotrophy complementation system. Plasmid 107: 102476.

    PubMed  CAS  Google Scholar 

  71. Hashiro, S. and H. Yasueda (2018) Plasmid copy number mutation in repA gene encoding RepA replication initiator of cryptic plasmid pHM1519 in Corynebacterium glutamicum. Biosci. Biotechnol. Biochem. 82: 2212–2224.

    PubMed  CAS  Google Scholar 

  72. Choi, J. W., S. S. Yim, and K. J. Jeong (2018) Development of a high-copy-number plasmid via adaptive laboratory evolution of Corynebacterium glutamicum. Appl. Microbiol. Biotechnol. 102: 873–883.

    PubMed  CAS  Google Scholar 

  73. Henke, N. A., I. Krahn, and V. F. Wendisch (2021) Improved plasmid-based inducible and constitutive gene expression in Corynebacterium glutamicum. Microorganisms 9: 204.

    PubMed  CAS  PubMed Central  Google Scholar 

  74. Jorge, J. M. P., F. Pérez-García, and V. F. Wendisch (2017) A new metabolic route for the fermentative production of 5-aminovalerate from glucose and alternative carbon sources. Bioresour. Technol. 245: 1701–1709.

    PubMed  CAS  Google Scholar 

  75. Kang, M. K., J. Lee, Y. Um, T. S. Lee, M. Bott, S. J. Park, and H. M. Woo (2014) Synthetic biology platform of CoryneBrick vectors for gene expression in Corynebacterium glutamicum and its application to xylose utilization. Appl. Microbiol. Biotechnol. 98: 5991–6002.

    PubMed  CAS  Google Scholar 

  76. Cleto, S., J. V. K. Jensen, V. F. Wendisch, and T. K. Lu (2016) Corynebacterium glutamicum metabolic engineering with CRISPR Interference (CRISPRi). ACS Synth. Biol. 5: 375–385.

    PubMed  CAS  PubMed Central  Google Scholar 

  77. Park, J., H. Shin, S.-M. Lee, Y. Um, and H. M. Woo (2018) RNA-guided single/double gene repressions in Corynebacterium glutamicum using an efficient CRISPR interference and its application to industrial strain. Microb. Cell Fact. 17: 4.

    PubMed  PubMed Central  Google Scholar 

  78. Yoon, J. and H. M. Woo (2018) CRISPR interference-mediated metabolic engineering of Corynebacterium glutamicum for homo-butyrate production. Biotechnol. Bioeng. 115: 2067–2074.

    PubMed  CAS  Google Scholar 

  79. Zhang, B., Z.-Q. Liu, C. Liu, and Y.-G. Zheng (2016) Application of CRISPRi in Corynebacterium glutamicum for shikimic acid production. Biotechnol. Lett. 38: 2153–2161.

    PubMed  CAS  Google Scholar 

  80. Gauttam, R., G. M. Seibold, P. Mueller, T. Weil, T. Weiß, R. Handrick, and B. J. Eikmanns (2019) A simple dual-inducible CRISPR interference system for multiple gene targeting in Corynebacterium glutamicum. Plasmid 103: 25–35.

    PubMed  CAS  Google Scholar 

  81. Yao, C., X. Hu, and X. Wang (2021) Construction and application of a CRISPR/Cas9-assisted genomic editing system for Corynebacterium glutamicum. AMB Express 11: 70.

    PubMed  CAS  PubMed Central  Google Scholar 

  82. Li, M., J. Chen, Y. Wang, J. Liu, J. Huang, N. Chen, P. Zheng, and J. Sun (2020) Efficient multiplex gene repression by CRISPR-dCpf1 in Corynebacterium glutamicum. Front. Bioeng. Biotechnol. 8: 357.

    PubMed  PubMed Central  Google Scholar 

  83. Calero, P. and P. I. Nikel (2019) Chasing bacterial chassis for metabolic engineering: a perspective review from classical to non-traditional microorganisms. Microb. Biotechnol. 12: 98–124.

    PubMed  CAS  Google Scholar 

  84. Unthan, S., M. Baumgart, A. Radek, M. Herbst, D. Siebert, N. Brühl, A. Bartsch, M. Bott, W. Wiechert, K. Marin, S. Hans, R. Krämer, G. Seibold, J. Frunzke, J. Kalinowski, C. Rückert, V. F. Wendisch, and S. Noack (2015) Chassis organism from Corynebacterium glutamicum—a top-down approach to identify and delete irrelevant gene clusters. Biotechnol. J. 10: 290–301.

    PubMed  CAS  Google Scholar 

  85. Choi, J. W., S. S. Yim, M. J. Kim, and K. J. Jeong (2015) Enhanced production of recombinant proteins with Corynebacterium glutamicum by deletion of insertion sequences (IS elements). Microb. Cell Fact. 14: 207.

    PubMed  PubMed Central  Google Scholar 

  86. Baumgart, M., S. Unthan, C. Rückert, J. Sivalingam, A. Grünberger, J. Kalinowski, M. Bott, S. Noack, and J. Frunzke (2013) Construction of a prophage-free variant of Corynebacterium glutamicum ATCC 13032 for use as a platform strain for basic research and industrial biotechnology. Appl. Environ. Microbiol. 79: 6006–6015.

    PubMed  CAS  PubMed Central  Google Scholar 

  87. Linder, M., M. Haak, A. Botes, J. Kalinowski, and C. Rückert (2021) Construction of an IS-free Corynebacterium glutamicum ATCC13032 chassis strain and random mutagenesis using the endogenous ISCg1 transposase. Front. Bioeng. Biotechnol. 9: 751334.

    PubMed  PubMed Central  Google Scholar 

  88. Baumgart, M., S. Unthan, R. Kloß, A. Radek, T. Polen, N. Tenhaef, M. F. Müller, A. Küberl, D. Siebert, N. Brühl, K. Marin, S. Hans, R. Krämer, M. Bott, J. Kalinowski, W. Wiechert, G. Seibold, J. Frunzke, C. Rückert, V. F. Wendisch, and S. Noack (2018) Corynebacterium glutamicum chassis C1*: building and testing a novel platform host for synthetic biology and industrial biotechnology. ACS Synth. Biol. 7: 132–144.

    PubMed  CAS  Google Scholar 

  89. Ferrer, L., M. Mindt, M. Suarez-Diez, T. Jilg, M. Zagorščak, J.-H. Lee, K. Gruden, V. F. Wendisch, and K. Cankar (2022) Fermentative indole production via bacterial tryptophan synthase alpha subunit and plant Indole-3-Glycerol phosphate lyase enzymes. J. Agric. Food Chem. 70: 5634–5645.

    PubMed  CAS  PubMed Central  Google Scholar 

  90. Walter, T., N. Al Medani, A. Burgardt, K. Cankar, L. Ferrer, A. Kerbs, J. H. Lee, M. Mindt, J. M. Risse, and V. F. Wendisch (2020) Fermentative N-methylanthranilate production by engineered Corynebacterium glutamicum. Microorganisms 8: 866.

    PubMed  CAS  PubMed Central  Google Scholar 

  91. Tsirigotaki, A., J. De Geyter, N. Šoštarić, A. Economou, and S. Karamanou (2017) Protein export through the bacterial Sec pathway. Nat. Rev. Microbiol. 15: 21–36.

    PubMed  CAS  Google Scholar 

  92. Brüser, T. (2007) The twin-arginine translocation system and its capability for protein secretion in biotechnological protein production. Appl. Microbiol. Biotechnol. 76: 35–45.

    PubMed  Google Scholar 

  93. Goosens, V. J., A. De-San-Eustaquio-Campillo, R. Carballido-López, and J. M. van Dijl (2015) A Tat ménage à trois—the role of Bacillus subtilis TatAc in twin-arginine protein translocation. Biochim. Biophys. Acta 1853: 2745–2753.

    PubMed  CAS  Google Scholar 

  94. Wang, Y., X. Gao, X. Liu, Y. Li, M. Sun, Y. Yang, C. Liu, and Z. Bai (2020) Construction of a 3A system from BioBrick parts for expression of recombinant hirudin variants III in Corynebacterium glutamicum. Appl. Microbiol. Biotechnol. 104: 8257–8266.

    PubMed  CAS  Google Scholar 

  95. Yu, X., X. Liu, X. Gao, X. Luo, Y. Yang, Y. Li, C. Liu, C. Zhang, and Z. Bai (2022) Development of a novel platform for recombinant protein production in Corynebacterium glutamicum on ethanol. Synth. Syst. Biotechnol. 7: 765–774.

    PubMed  PubMed Central  Google Scholar 

  96. Sun, M., A. X. Gao, R. Ledesma-Amaro, A. Li, R. Wang, J. Nie, P. Zheng, Y. Yang, Z. Bai, and X. Liu (2022) Hypersecretion of OmlA antigen in Corynebacterium glutamicum through high-throughput based development process. Appl. Microbiol. Biotechnol. 106: 2953–2967.

    PubMed  CAS  Google Scholar 

  97. Kikuchi, Y., H. Itaya, M. Date, K. Matsui, and L.-F. Wu (2009) TatABC overexpression improves Corynebacterium glutamicum Tat-dependent protein secretion. Appl. Environ. Microbiol. 75: 603–607.

    PubMed  CAS  Google Scholar 

  98. Teramoto, H., K. Watanabe, N. Suzuki, M. Inui, and H. Yukawa (2011) High yield secretion of heterologous proteins in Corynebacterium glutamicum using its own Tat-type signal sequence. Appl. Microbiol. Biotechnol. 91: 677–687.

    PubMed  CAS  Google Scholar 

  99. Scheele, S., D. Oertel, J. Bongaerts, S. Evers, H. Hellmuth, K.-H. Maurer, M. Bott, and R. Freudl (2013) Secretory production of an FAD cofactor-containing cytosolic enzyme (sorbitol-xylitol oxidase from Streptomyces coelicolor) using the twin-arginine translocation (Tat) pathway of Corynebacterium glutamicum. Microb. Biotechnol. 6: 202–206.

    PubMed  Google Scholar 

  100. Matano, C., S. Kolkenbrock, S. N. Hamer, E. Sgobba, B. M. Moerschbacher, and V. F. Wendisch (2016) Corynebacterium glutamicum possesses ß-N-acetylglucosaminidase. BMC Microbiol. 16: 177.

    PubMed  PubMed Central  Google Scholar 

  101. Jeon, E. J., J. W. Choi, M. S. Cho, and K. J. Jeong (2021) Enhanced production of neoagarobiose from agar with Corynebacterium glutamicum producing exo-type and endotype β-agarases. Microb. Biotechnol. 14: 2164–2175.

    PubMed  CAS  PubMed Central  Google Scholar 

  102. Yim, S. S., J. W. Choi, R. J. Lee, Y. J. Lee, S. H. Lee, S. Y. Kim, and K. J. Jeong (2016) Development of a new platform for secretory production of recombinant proteins in Corynebacterium glutamicum. Biotechnol. Bioeng. 113: 163–172.

    PubMed  CAS  Google Scholar 

  103. Watanabe, K., Y. Tsuchida, N. Okibe, H. Teramoto, N. Suzuki, M. Inui, and H. Yukawa (2009) Scanning the Corynebacterium glutamicum R genome for high-efficiency secretion signal sequences. Microbiology (Reading) 155: 741–750.

    PubMed  CAS  Google Scholar 

  104. Hemmerich, J., P. Rohe, B. Kleine, S. Jurischka, W. Wiechert, R. Freudl, and M. Oldiges (2016) Use of a Sec signal peptide library from Bacillus subtilis for the optimization of cutinase secretion in Corynebacterium glutamicum. Microb. Cell Fact. 15: 208.

    PubMed  PubMed Central  Google Scholar 

  105. Westers, H., L. Westers, E. Darmon, J. M. van Dijl, W. J. Quax, and G. Zanen (2006) The CssRS two-component regulatory system controls a general secretion stress response in Bacillus subtilis. FEBS J. 273: 3816–3827.

    PubMed  CAS  Google Scholar 

  106. Jurischka, S., A. Bida, D. Dohmen-Olma, B. Kleine, J. Potzkei, S. Binder, G. Schaumann, P. J. Bakkes, and R. Freudl (2020) A secretion biosensor for monitoring Sec-dependent protein export in Corynebacterium glutamicum. Microb. Cell Fact. 19: 11.

    PubMed  CAS  PubMed Central  Google Scholar 

  107. Knapp, A., M. Ripphahn, K. Volkenborn, P. Skoczinski, and K.-E. Jaeger (2017) Activity-independent screening of secreted proteins using split GFP. J. Biotechnol. 258: 110–116.

    PubMed  CAS  Google Scholar 

  108. Bakkes, P. J., P. Lenz, C. Müller, A. Bida, D. Dohmen-Olma, A. Knapp, M. Oldiges, K.-E. Jaeger, and R. Freudl (2021) Biosensor-based optimization of cutinase secretion by Corynebacterium glutamicum. Front. Microbiol. 12: 750150.

    PubMed  PubMed Central  Google Scholar 

  109. Cabantous, S., T. C. Terwilliger, and G. S. Waldo (2005) Protein tagging and detection with engineered self-assembling fragments of green fluorescent protein. Nat. Biotechnol. 23: 102–107.

    PubMed  CAS  Google Scholar 

  110. Wang, B. L., A. Ghaderi, H. Zhou, J. Agresti, D. A. Weitz, G. R. Fink, and G. Stephanopoulos (2014) Microfluidic high-throughput culturing of single cells for selection based on extracellular metabolite production or consumption. Nat. Biotechnol. 32: 473–478.

    PubMed  CAS  PubMed Central  Google Scholar 

  111. Chen, J., M. Vestergaard, T. G. Jensen, J. Shen, M. Dufva, C. Solem, and P. R. Jensen (2017) Finding the needle in the haystack-the use of microfluidic droplet technology to identify vitamin-secreting lactic acid bacteria. mBio 8: e00526–17.

    PubMed  CAS  PubMed Central  Google Scholar 

  112. Balasubramanian, S., J. Chen, V. Wigneswaran, C. H. Bang-Berthelsen, and P. R. Jensen (2021) Droplet-based microfluidic high throughput screening of Corynebacterium glutamicum for efficient heterologous protein production and secretion. Front. Bioeng. Biotechnol. 9: 668513.

    PubMed  PubMed Central  Google Scholar 

  113. Bayan, N., C. Houssin, M. Chami, and G. Leblon (2003) Mycomembrane and S-layer: two important structures of Corynebacterium glutamicum cell envelope with promising biotechnology applications. J. Biotechnol. 104: 55–67.

    PubMed  CAS  Google Scholar 

  114. Matsuda, Y., H. Itaya, Y. Kitahara, N. M. Theresia, E. A. Kutukova, Y. A. V. Yomantas, M. Date, Y. Kikuchi, and M. Wachi (2014) Double mutation of cell wall proteins CspB and PBP1a increases secretion of the antibody Fab fragment from Corynebacterium glutamicum. Microb. Cell Fact. 13: 56.

    PubMed  PubMed Central  Google Scholar 

  115. Jin, Q., F. Pan, C.-F. Hu, S. Y. Lee, X.-X. Xia, and Z.-G. Qian (2022) Secretory production of spider silk proteins in metabolically engineered Corynebacterium glutamicum for spinning into tough fibers. Metab. Eng. 70: 102–114.

    PubMed  CAS  Google Scholar 

  116. Hemmerich, J., M. Labib, C. Steffens, S. J. Reich, M. Weiske, M. Baumgart, C. Rückert, M. Ruwe, D. Siebert, V. F. Wendisch, J. Kalinowski, W. Wiechert, and M. Oldiges (2020) Screening of a genome-reduced Corynebacterium glutamicum strain library for improved heterologous cutinase secretion. Microb. Biotechnol. 13: 2020–2031.

    PubMed  CAS  PubMed Central  Google Scholar 

  117. Overton, T. W. (2014) Recombinant protein production in bacterial hosts. Drug Discov. Today 19: 590–601.

    PubMed  CAS  Google Scholar 

  118. Ryan, B. J. and G. T. Henehan (2013) Overview of approaches to preventing and avoiding proteolysis during expression and purification of proteins. Curr. Protoc. Protein Sci. Chapter 5: Unit5.25.

  119. Lüdke, A., R. Krämer, A. Burkovski, D. Schluesener, and A. Poetsch (2007) A proteomic study of Corynebacterium glutamicum AAA+ protease FtsH. BMC Microbiol. 7: 6.

    PubMed  PubMed Central  Google Scholar 

  120. Hong, E. J., J. S. Park, Y. Kim, and H. S. Lee (2014) Role of Corynebacterium glutamicum sprA encoding a serine protease in glxR-mediated global gene regulation. PLoS One 9: e93587.

    PubMed  PubMed Central  Google Scholar 

  121. Engels, S., J. E. Schweitzer, C. Ludwig, M. Bott, and S. Schaffer (2004) clpC and clpP1P2 gene expression in Corynebacterium glutamicum is controlled by a regulatory network involving the transcriptional regulators ClgR and HspR as well as the ECF sigma factor sigmaH. Mol. Microbiol. 52: 285–302.

    PubMed  CAS  Google Scholar 

  122. Liu, X., L. Meng, X. Wang, Y. Yang, and Z. Bai (2022) Effect of Clp protease from Corynebacterium glutamicum on heterologous protein expression. Protein Expr. Purif. 189: 105928.

    PubMed  CAS  Google Scholar 

  123. Peng, F., X. Liu, X. Wang, J. Chen, M. Liu, Y. Yang, and Z. Bai (2019) Triple deletion of clpC, porB, and mepA enhances production of small ubiquitin-like modifier-N-terminal probrain natriuretic peptide in Corynebacterium glutamicum. J. Ind. Microbiol. Biotechnol. 46: 67–79.

    PubMed  CAS  Google Scholar 

  124. Park, J., S. Lee, M. J. Lee, K. Park, S. Lee, J. F. Kim, and P. Kim (2020) Accelerated growth of Corynebacterium glutamicum by up-regulating stress- responsive genes based on transcriptome analysis of a fast-doubling evolved strain. J. Microbiol. Biotechnol. 30: 1420–1429.

    PubMed  CAS  PubMed Central  Google Scholar 

  125. Graf, M., T. Haas, F. Müller, A. Buchmann, J. Harm-Bekbenbetova, A. Freund, A. Nieß, M. Persicke, J. Kalinowski, B. Blombach, and R. Takors (2019) Continuous adaptive evolution of a fast-growing Corynebacterium glutamicum strain independent of protocatechuate. Front. Microbiol. 10: 1648.

    PubMed  PubMed Central  Google Scholar 

  126. Paalme, T., R. Elken, A. Kahru, K. Vanatalu, and R. Vilu (1997) The growth rate control in Escherichia coli at near to maximum growth rates: the A-stat approach. Antonie Van Leeuwenhoek 71: 217–230.

    PubMed  CAS  Google Scholar 

  127. Pfeifer, E., C. Gätgens, T. Polen, and J. Frunzke (2017) Adaptive laboratory evolution of Corynebacterium glutamicum towards higher growth rates on glucose minimal medium. Sci. Rep. 7: 16780.

    PubMed  PubMed Central  Google Scholar 

  128. Lee, M. J., J. Park, K. Park, J. F. Kim, and P. Kim (2020) Reverse engineering targets for recombinant protein production in Corynebacterium glutamicum inspired by a fast-growing evolved descendant. Front. Bioeng. Biotechnol. 8: 588070.

    PubMed  PubMed Central  Google Scholar 

  129. Parwin, S., S. Kalan, and P. Srivastava (2019) Bacterial cell surface display. pp. 81–108. In: N. K. Rathinam and R. K. Sani (eds.). Next Generation Biomanufacturing Technologies. American Chemical Society, Washington, DC, USA.

    Google Scholar 

  130. Kim, D., W. Kim, and J. Kim (2021) New bacterial surface display system development and application based on Bacillus subtilis YuaB biofilm component as an anchoring motif. Biotechnol. Bioprocess Eng. 26: 39–46.

    PubMed  CAS  PubMed Central  Google Scholar 

  131. Tateno, T., H. Fukuda, and A. Kondo (2007) Production of L-Lysine from starch by Corynebacterium glutamicum displaying alpha-amylase on its cell surface. Appl. Microbiol. Biotechnol. 74: 1213–1220.

    PubMed  CAS  Google Scholar 

  132. Tateno, T., K. Hatada, T. Tanaka, H. Fukuda, and A. Kondo (2009) Development of novel cell surface display in Corynebacterium glutamicum using porin. Appl. Microbiol. Biotechnol. 84: 733–739.

    PubMed  CAS  Google Scholar 

  133. Choi, J. W., S. S. Yim, and K. J. Jeong (2018) Development of a potential protein display platform in Corynebacterium glutamicum using mycolic acid layer protein, NCgl1337, as an anchoring motif. Biotechnol. J. 13: 1700509.

    Google Scholar 

  134. Lin, K., N. Zhao, Y. Cai, Y. Lin, S. Han, and S. Zheng (2022) Genome-scale mining of novel anchor proteins of Corynebacterium glutamicum. Front. Microbiol. 12: 677702.

    PubMed  PubMed Central  Google Scholar 

  135. Koo, B. I., J. W. Choi, S. Y. Song, Y. H. Choi, T. Y. Lee, S.-H. Kim, K. J. Jeong, and Y. S. Nam (2021) Robust biocatalysts displayed on crystalline protein-layered cells for efficient and sustainable hydration of carbon dioxide. Adv. Funct. Mater. 31:2102497.

    CAS  Google Scholar 

  136. Jin, C., J. Li, Z. Huang, X. Han, and J. Bao (2022) Engineering Corynebacterium glutamicum for synthesis of poly(3-hydroxy-butyrate) from lignocellulose biomass. Biotechnol. Bioeng. 119: 1598–1613.

    PubMed  CAS  Google Scholar 

  137. Adachi, N., C. Takahashi, N. Ono-Murota, R. Yamaguchi, T. Tanaka, and A. Kondo (2013) Direct L-lysine production from cellobiose by Corynebacterium glutamicum displaying beta-glucosidase on its cell surface. Appl. Microbiol. Biotechnol. 97: 7165–7172.

    PubMed  CAS  Google Scholar 

  138. Kim, S. J., J. E. Hyeon, S. D. Jeon, G. W. Choi, and S. O. Han (2014) Bi-functional cellulases complexes displayed on the cell surface of Corynebacterium glutamicum increase hydrolysis of lignocelluloses at elevated temperature. Enzyme Microb. Technol. 66: 67–73.

    PubMed  CAS  Google Scholar 

  139. Buchholz, J., A. Schwentner, B. Brunnenkan, C. Gabris, S. Grimm, R. Gerstmeir, R. Takors, B. J. Eikmanns, and B. Blombach (2013) Platform engineering of Corynebacterium glutamicum with reduced pyruvate dehydrogenase complex activity for improved production of L-lysine, L-valine, and 2-ketoisovalerate. Appl. Environ. Microbiol. 79: 5566–5575.

    PubMed  CAS  PubMed Central  Google Scholar 

  140. Ravasi, P., M. Braia, F. Eberhardt, C. Elena, S. Cerminati, S. Peirú, M. E. Castelli, and H. G. Menzella (2015) High-level production of Bacillus cereus phospholipase C in Corynebacterium glutamicum. J. Biotechnol. 216: 142–148.

    PubMed  CAS  Google Scholar 

  141. Zhang, J., S. D. Petersen, T. Radivojevic, A. Ramirez, A. Pérez-Manríquez, E. Abeliuk, B. J. Sánchez, Z. Costello, Y. Chen, M. J. Fero, H. G. Martin, J. Nielsen, J. D. Keasling, and M. K. Jensen (2020) Combining mechanistic and machine learning models for predictive engineering and optimization of tryptophan metabolism. Nat. Commun. 11: 4880.

    PubMed  PubMed Central  Google Scholar 

  142. Oyetunde, T., F. S. Bao, J.-W. Chen, H. G. Martin, and Y. J. Tang (2018) Leveraging knowledge engineering and machine learning for microbial bio-manufacturing. Biotechnol. Adv. 36: 1308–1315.

    PubMed  CAS  Google Scholar 

  143. Kwon, M. S., B. T. Lee, S. Y. Lee, and H. U. Kim (2020) Modeling regulatory networks using machine learning for systems metabolic engineering. Curr. Opin. Biotechnol. 65: 163–170.

    PubMed  CAS  Google Scholar 

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Acknowledgements

This work was supported by Korea Institute of Planning and Evaluation for Technology in Food, Agriculture and Forestry (IPET) through Crop Viruses and Pests Response Industry Technology Development Program, funded by Ministry of Agriculture, Food and Rural Affairs (MAFRA) (Grant no. 321109-04-1-HD020), and by the Cooperative Research Program for Agriculture Science and Technology Development (project number PJ015613022021) of the Rural Development Administration, Republic of Korea.

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Lee, S.M., Jeong, K.J. Advances in Synthetic Biology Tools and Engineering of Corynebacterium glutamicum as a Platform Host for Recombinant Protein Production. Biotechnol Bioproc E 28, 962–976 (2023). https://doi.org/10.1007/s12257-022-0219-1

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