Skip to main content
SpringerLink
Log in

Purification and Biochemical Characterization of a Tyrosine Phenol-lyase from Morganella morganii

  • Published:
Applied Biochemistry and Biotechnology Aims and scope Submit manuscript

Abstract

Tyrosine phenol-lyase (TPL) is a valuable and cost-effective biocatalyst for the biosynthesis of L-tyrosine and its derivatives, which are valuable intermediates in the pharmaceutical industry. A TPL from Morganella morganii (Mm-TPL) was overexpressed in Escherichia coli and characterized. Mm-TPL was determined as a homotetramer with molecular weight of 52 kDa per subunit. Its optimal temperature and pH for β-elimination of L-tyrosine were 45 °C and pH 8.5, respectively. Mm-TPL manifested strict substrate specificity for the reverse reaction of β-elimination and ortho- and meta-substituted phenols with small steric size were preferred substrates. The enzyme showed excellent catalytic performance for synthesis of L-tyrosine, 3-fluoro-L-tyrosine, and L-DOPA with a yield of 98.1%, 95.1%, and 87.2%, respectively. Furthermore, the fed-batch bioprocess displayed space-time yields of 9.6 g L−1 h−1 for L-tyrosine and 4.2 g L−1 h−1 for 3-fluoro-L-tyrosine with a yield of 67.4 g L−1 and 29.5 g L−1, respectively. These results demonstrated the great potential of Mm-TPL for industrial application.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Scheme 1
Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6

Similar content being viewed by others

References

  1. Milic, D., Demidkina, T. V., Faleev, N. G., Phillips, R. S., Matkovic-Calogovic, D., & Antson, A. A. (2011). Crystallographic snapshots of tyrosine phenol-lyase show that substrate strain plays a role in C-C bond cleavage. Journal of the American Chemical Society, 133(41), 16468–16476.

    CAS  PubMed  PubMed Central  Google Scholar 

  2. Yamada, H., & Kumagai, H. (1975). Synthesis of L-tyrosine-related amino acids by beta-tyrosinase. Advances in Applied Microbiology, 19, 249–288.

    CAS  PubMed  Google Scholar 

  3. Phillips, R. S., Chen, H. Y., & Faleev, N. G. (2006). Aminoacrylate intermediates in the reaction of Citrobacter freundii tyrosine phenol-lyase. Biochemistry, 45(31), 9575–9583.

    CAS  PubMed  Google Scholar 

  4. Santos, C. N. S., & Stephanopoulos, G. (2007). Melanin-based high-throughput screen for L-tyrosine production in Escherichia coli. Applied and Environmental Microbiology, 74(4), 1190–1197.

    PubMed  PubMed Central  Google Scholar 

  5. Santos, C. N. S., Xiao, W. H., & Stephanopoulos, G. (2012). Rational, combinatorial, and genomic approaches for engineering L-tyrosine production in Escherichia coli. Proceedings of the National Academy of Sciences, 109(34), 13538–13543.

    CAS  Google Scholar 

  6. Patil, S. A., Apine, O. A., Surwase, S. N., & Jadhav, J. P. (2013). Biological sources of L-DOPA: an alternative approach. Advances in Parkinson's Disease, 02(03), 81–87.

    Google Scholar 

  7. Monclus, M., Masson, C., & Luxen, A. (1995). Asymmetric synthesis of fluorinated L-tyrosine and meta-L-tyrosines. Journal of Fluorine Chemistry, 70, 39–43.

    CAS  Google Scholar 

  8. Reid, P. J., Loftus, C., Beeson, C. C., & Study, U. V. r. R. (2003). Evaluating the potential of fluorinated tyrosines as spectroscopic probes of local protein environments: a UV resonance Raman study. Biochemistry, 42(8), 2441–2448.

    CAS  PubMed  Google Scholar 

  9. Ikeda, M. (2002). Amino acid production processes. Advances in Biochemical Engineering/Biotechnology, 79, 1–35.

    Google Scholar 

  10. Noisier, A. F., Harris, C. S., & Brimble, M. A. (2013). Novel preparation of chiral alpha-amino acids using the Mitsunobu-Tsunoda reaction. Chemical Communications, 49(70), 7744–7746.

    CAS  PubMed  Google Scholar 

  11. Juminaga, D., Baidoo, E. E. K., Redding-Johanson, A. M., Batth, T. S., Burd, H., Mukhopadhyay, A., Petzold, C. J., & Keasling, J. D. (2012). Modular engineering of L-tyrosine production in Escherichia coli. Applied and Environmental Microbiology, 78(1), 89–98.

    CAS  PubMed  PubMed Central  Google Scholar 

  12. Xue, Y. P., Cao, C. H., & Zheng, Y. G. (2018). Enzymatic asymmetric synthesis of chiral amino acids. Chemical Society Reviews, 47(4), 1516–1561.

    CAS  PubMed  Google Scholar 

  13. Milic, D., Matkovic-Calogovic, D., Demidkina, T. V., Kulikova, V. V., Sinitzina, N. I., & Antson, A. A. (2006). Structures of apo- and holo-tyrosine phenol-lyase reveal a catalytically critical closed conformation and suggest a mechanism for activation by K+ ions. Biochemistry, 45(24), 7544–7552.

    CAS  PubMed  PubMed Central  Google Scholar 

  14. Ogawa, J., & Shimizu, S. (2002). Industrial microbial enzymes: their discovery by screening and use in large-scale production of useful chemicals in Japan. Current Opinion in Biotechnology, 13(4), 367–375.

    CAS  PubMed  Google Scholar 

  15. Tang, X. L., Suo, H., Wang, Z. C., Zheng, R. C., & Zheng, Y. G. (2018). Process development for efficient biosynthesis of L-DOPA with recombinant Escherichia coli harboring tyrosine phenol lyase from Fusobacterium nucleatum. Bioprocess and Biosystems Engineering, 41(9), 1347–1354.

    CAS  PubMed  Google Scholar 

  16. Kim, D. Y., Rha, E., Choi, S. L., Song, J. J., Hong, S. P., Sung, M. H., & Lee, S. G. (2007). Development of bioreactor system for L-tyrosine synthesis using thermostable tyrosine phenol-lyase. Journal of Microbiology and Biotechnology, 17(1), 116–122.

    CAS  PubMed  Google Scholar 

  17. Lozinsky, V. I., Faleev, N. G., Zubov, A. L., Ruvinov, S. B., Antonova, T. V., Vainerman, E. S., Belikov, V. M., & Rogozhin, S. V. (1989). Use of PVA-cryogel entrapped Citrobacter intermedius cells for continuous production of 3-fluoro-L-tyrosine. Biotechnology Letters, 11(1), 43–48.

    Article  Google Scholar 

  18. Zhang, H. J., Lu, Y., Wu, S. P., Wei, Y., Liu, Q., Liu, J. Z., & Jiao, Q. C. (2016). Two-step enzymatic synthesis of tyramine from raw pyruvate fermentation broth. Journal of Molecular Catalysis B: Enzymatic, 124, 38–44.

    Article  CAS  Google Scholar 

  19. Kumagai, H., Kashima, N., & Yamada, H. (1970). Racemization of D- or L-alanine by crystalline tyrosine phenol-lyase from Eshherichia intermedia. Biochemical and Biophysical Research Communications, 39(5), 796–801.

    Article  CAS  PubMed  Google Scholar 

  20. Chandel, M., & Azmi, W. (2013). Purification and characterization of tyrosine phenol lyase from Citrobacter freundii. Applied Biochemistry and Biotechnology, 171(8), 2040–2052.

    CAS  PubMed  Google Scholar 

  21. Kumagai, H., Kashima, N., Torii, H., Yamada, H., Enei, H., & Okumuea, S. (1971). Purification, crystallization and properties of tyrosine phenol lyase from Erwinia herbicola. Agricultural and Biological Chemistry, 36(3), 472–482.

    Google Scholar 

  22. Rhee, S. K., Lee, S. G., Hong, S. P., Choi, Y. H., Park, J. H., Kim, C. J., & Sung, M. H. (2000). A novel microbial interaction: obligate commensalism between a new gram-negative thermophile and a thermophilic Bacillus strain. Extremophiles, 4(3), 131–136.

    CAS  PubMed  Google Scholar 

  23. Zheng, R. C., Tang, X. L., Suo, H., Feng, L. L., Liu, X., Yang, J., & Zheng, Y. G. (2018). Biochemical characterization of a novel tyrosine phenol-lyase from Fusobacterium nucleatum for highly efficient biosynthesis of l-DOPA. Enzyme and Microbial Technology, 112, 88–93.

    CAS  PubMed  Google Scholar 

  24. Wynands, B., Lenzen, C., Otto, M., Koch, F., Blank, L. M., & Wierckx, N. (2018). Metabolic engineering of Pseudomonas taiwanensis VLB120 with minimal genomic modifications for high-yield phenol production. Metabolic Engineering, 47, 121–133.

    CAS  PubMed  Google Scholar 

  25. Carman, G. M., & Levin, R. E. (1977). Partial purification and some properties of tyrosine phenol-lyase from Aeromonas phenologenes ATCC 29063. Applied and Environmental Microbiology, 33(1), 192–198.

    CAS  PubMed  PubMed Central  Google Scholar 

  26. Brot, N., Smit, Z., & Weissbach, H. (1965). Conversion of l-tyrosine to phenol by Clostridium tetanomorphum. Archives of Biochemistry and Biophysics, 112(1), 1–6.

    CAS  PubMed  Google Scholar 

  27. Enei, H., Nakazawa, H., Okumura, S., & Yamada, H. (1973). Synthesis of L-tyrosine or 3, 4-dihydroxyphenyl-L-alanine from pyruvic acid, ammonia and phenol or pyrocatechol. Agricultural and Biological Chemistry, 37(4), 725–735.

    CAS  Google Scholar 

  28. Milic, D., Demidkina, T. V., Faleev, N. G., Matkovic-Calogovic, D., & Antson, A. A. (2008). Insights into the catalytic mechanism of tyrosine phenol-lyase from X-ray structures of quinonoid intermediates. The Journal of Biological Chemistry, 283(43), 29206–29214.

    CAS  PubMed  PubMed Central  Google Scholar 

  29. Faleev, N. G., Axenova, O. V., Demidkina, T. V., & Phillips, R. S. (2003). The role of acidic dissociation of substrate’s phenol group in the mechanism of tyrosine phenol-lyase. Biochimica et Biophysica Acta, 1647(1-2), 260–265.

    CAS  PubMed  Google Scholar 

  30. Lee, S. G., Hong, S. P., Kwak, M. S., Esaki, N., & Sung, M. H. (1999). Characterization of thermostable tyrosine phenol-lyase from an obligatory symbiotic thermophile, Symbiobacterium sp. SC-1. Journal of Biochemistry and Molecular Biology, 32(5), 480–485.

    CAS  Google Scholar 

  31. Kumagai, H., Yamada, H., Matsui, H., Ohkishi, H., & Ogata, K. (1970). Tyrosine phenol lyase. I. Purification, crystallization, and properties. The Journal of Biological Chemistry, 245(7), 1767–1772.

    CAS  PubMed  Google Scholar 

  32. Miagkikh, I. V., & Demidkina, T. V. (1985). Effect of monovalent cations on the catalytic and spectral properties of tyrosine-phenol-lyase from Citrobacter intermedius. Molecular Biology, 19(3), 671–678.

    CAS  Google Scholar 

  33. Sundararaju, B., Chen, H., Shilcutt, S., & Phillips, R. S. (2000). The role of glutamic acid-69 in the activation of Citrobacter freundii tyrosine phenol-lyase by monovalent cations. Biochemistry, 39(29), 8546–8555.

    CAS  PubMed  Google Scholar 

  34. Nagasawa, T., Utagawa, T., Goto, J., Kim, C. J., Tani, Y., Kumagai, H., & Yamada, H. (1981). Syntheses of L-tyrosine-related amino acids by tyrosine phenol-lyase of Citrobacter intermedius. European Journal of Biochemistry, 117(1), 33–40.

    CAS  PubMed  Google Scholar 

  35. Hay, P. J., & Shavitt, I. (1974). Ab initioconfiguration interaction studies of the π-electron states of benzene. The Journal of Chemical Physics, 60(7), 2865–2877.

    CAS  Google Scholar 

  36. Skalický, T., Chollet, C., Pasquier, N., & Allan, M. (2002). Properties of the π* and σ* states of the chlorobenzene anion determined by electron impact spectroscopy. Physical Chemistry Chemical Physics, 4(15), 3583–3590.

    Google Scholar 

  37. Phillips, R. S., Von Tersch, R. L., & Secundo, F. (1997). Effects of tyrosine ring fluorination on rates and equilibria of formation of intermediates in the reactions of carbon-carbon lyases. European Journal of Biochemistry, 244(2), 658–663.

    CAS  PubMed  Google Scholar 

  38. Tina, L. E., Santos, C. N. S., & Stephanopoulos, G. (2007). Perspectives of biotechnological production of L-tyrosine and its applications. Applied Microbiology and Biotechnology, 77(4), 751–762.

    Google Scholar 

Download references

Funding

This study was financially supported by the National Natural Science Foundation of China (No. 31900912) and Natural Science Foundation of Zhejiang Province (LQ17B06004).

Author information

Authors and Affiliations

  1. Key Laboratory of Bioorganic Synthesis of Zhejiang Province, College of Biotechnology and Bioengineering, Zhejiang University of Technology, Hangzhou, 310014, People’s Republic of China

    Hang-Qin Zhu, Xiao-Ling Tang, Ren-Chao Zheng & Yu-Guo Zheng

  2. Engineering Research Center of Bioconversion and Biopurification of Ministry of Education, Zhejiang University of Technology, Hangzhou, 310014, People’s Republic of China

    Hang-Qin Zhu, Xiao-Ling Tang, Ren-Chao Zheng & Yu-Guo Zheng

Authors
  1. Hang-Qin Zhu
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Xiao-Ling Tang
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Ren-Chao Zheng
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Yu-Guo Zheng
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Ren-Chao Zheng.

Ethics declarations

Conflict of Interest

The authors declare that they have no conflict of interest.

Additional information

Publisher’s Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Zhu, HQ., Tang, XL., Zheng, RC. et al. Purification and Biochemical Characterization of a Tyrosine Phenol-lyase from Morganella morganii. Appl Biochem Biotechnol 192, 71–84 (2020). https://doi.org/10.1007/s12010-020-03301-1

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12010-020-03301-1

Keywords

Search

Navigation