Advertisement

Applied Biochemistry and Biotechnology

, Volume 187, Issue 1, pp 75–89 | Cite as

One-Pot Enzymatic Synthesis of d-Arylalanines Using Phenylalanine Ammonia Lyase and l-Amino Acid Deaminase

  • Longbao Zhu
  • Guoqiang Feng
  • Fei Ge
  • Ping Song
  • Taotao Wang
  • Yi LiuEmail author
  • Yugui Tao
  • Zhemin Zhou
Article

Abstract

The phenylalanine ammonia-lyase (AvPAL) from Anabaena variabilis catalyzes the amination of substituent trans-cinnamic acid (t-CA) to produce racemic d,l-enantiomer arylalanine mixture owing to its low stereoselectivity. To produce high optically pure d-arylalanine, a modified AvPAL with high d-selectivity is expected. Based on the analyses of catalytic mechanism and structure, the Asn347 residue in the active site was proposed to control stereoselectivity. Therefore, Asn347 was mutated to construct mutant AvPAL-N347A, the stereoselectivity of AvPAL-N347A for d-enantiomer arylalanine was 2.3-fold higher than that of wild-type AvPAL (WtPAL). Furthermore, the residual l-enantiomer product in reaction solution could be converted into the d-enantiomer product through stereoselective oxidation by PmLAAD and nonselective reduction by reducing agent NH3BH3. At optimal conditions, the conversion rate of t-CA and optical purity (enantiomeric excess (eeD)) of d-phenylalanine reached 82% and exceeded 99%, respectively. The two enzymes displayed activity toward a broad range of substrate and could be used to efficiently synthesize d-arylalanine with different groups on the phenyl ring. Among these d-arylalanines, the yield of m-nitro-d-phenylalanine was highest and reached 96%, and the eeD exceeded 99%. This one-pot synthesis using AvPAL and PmLAAD has prospects for industrial application.

Keywords

Phenylalanine ammonia lyase l-Amino acid deaminase d-Arylalanine One-pot synthesis Gene cloning and expression Mutation 

Notes

Acknowledgments

The authors gratefully acknowledged Prof. Zheming Zhou from the School of Biotechnology, Jiangnan University for the facilities and infrastructure. The authors also acknowledged the help rendered by Weifeng Sun from Xihua University in proofreading the manuscript.

Funding

This work was supported by the National Natural Science Foundation of China (21506172; 31671797), Natural Sciences Foundation supported by Anhui Province universities (KJ2016A801) and Anhui Polytechnic University Youth Talent Support Program (2016BJRC006).

Compliance with Ethical Standards

Conflict of Interest

The authors declare that they have no conflict of interest.

Supplementary material

12010_2018_2794_MOESM1_ESM.doc (48 kb)
ESM 1 (DOC 48 kb)

References

  1. 1.
    Genchi, G. (2017). An overview on D-amino acids. Amino Acids, 49(9), 1521–1533.Google Scholar
  2. 2.
    Parmeggiani, F., Ahmed, S. T., Thompson, M. P., Weise, N. J., Galman, J. L., Gahloth, D., Dunstan, M. S., Leys, D., & Turner, N. J. (2016). Single-biocatalyst synthesis of enantiopure D-arylalanines exploiting an engineered D-amino acid dehydrogenase. Advanced Synthesis and Catalysis, 358(20), 3298–3306.Google Scholar
  3. 3.
    Fox, M. E., Jackson, M., Meek, G., & Willets, M. (2011). Large-scale synthesis of a substituted D-phenylalanine using asymmetric hydrogenation. Organic Process Research and Development, 15(5), 1163–1171.Google Scholar
  4. 4.
    Muller, U., van Assema, F., Gunsior, M., Orf, S., Kremer, S., Schipper, D., Wagemans, A., Townsend, C. A., Sonke, T., & Bovenberg, R. (2006). Metabolic engineering of the E. coli L-phenylalanine pathway for the production of D-phenylglycine (D-Phg). Metabolic Engineering, 8(3), 196–208.Google Scholar
  5. 5.
    Breuer, M., Ditrich, K., Habicher, T., Hauer, B., Kesseler, M., Sturmer, R., & Zelinski, T. (2004). Industrial methods for the production of optically active intermediates. Angewandte Chemie, International Edition, 43(7), 788–824.Google Scholar
  6. 6.
    Kobayashi, J., Shimizu, Y., Mutaguchi, Y., Doi, K., & Ohshima, T. (2013). Characterization of D-amino acid aminotransferase from Lactobacillus salivarius. Journal of Molecular Catalysis B Enzymatic, 94, 15–22.Google Scholar
  7. 7.
    Liu, R. X., Liu, S. P., Cheng, S., Zhang, L., Ding, Z. Y., Gu, Z. H., & Shi, G. Y. (2015). Screening, characterization and utilization of D-amino acid aminotransferase to obtain D-phenylalanine. Applied Biochemistry and Microbiology, 51(6), 695–703.Google Scholar
  8. 8.
    Hossain, G. S., Li, J., Shin, H., Du, G., Liu, L., & Chen, J. (2014). L-Amino acid oxidases from microbial sources: types, properties, functions, and applications. Applied Microbiology and Biotechnology, 98(4), 1507–1515.Google Scholar
  9. 9.
    Singh, S., Gogoi, B. K., & Bezbaruah, R. L. (2011). Racemic resolution of some DL-amino acids using Aspergillus fumigatus L-amino acid oxidase. Current Microbiology, 63(1), 94–99.Google Scholar
  10. 10.
    Park, J. H., Kim, G. J., & Kim, H. S. (2000). Production of D-amino acid using whole cells of recombinant Escherichia coli with separately and coexpressed D-hydantoinase and N-carbamoylase. Biotechnology Progress, 16(4), 564–570.Google Scholar
  11. 11.
    Altenbuchner, J., Siemann-Herzberg, M., & Syldatk, C. (2001). Hydantoinases and related enzymes as biocatalysts for the synthesis of unnatural chiral amino acids. Current Opinion in Biotechnology, 12(6), 559–563.Google Scholar
  12. 12.
    Gao, X. Z., Ma, Q. Y., & Zhu, H. L. (2015). Distribution, industrial applications, and enzymatic synthesis of D-amino acids. Applied Microbiology and Biotechnology, 99(8), 3341–3349.Google Scholar
  13. 13.
    Foster, I. M., Dorrington, R. D., & Burton, S. G. (2003). Enhanced hydantoinase and N-carbamoylase activity on immobilisation of Agrobacterium tumefaciens. Biotechnology Letters, 25(1), 67–72.Google Scholar
  14. 14.
    Parmeggiani, F., Ahmed, S. T., Weise, N. J., & Turner, N. J. (2016). Telescopic one-pot condensation-hydroamination strategy for the synthesis of optically pure L-phenylalanines from benzaldehydes. Tetrahedron, 72(46), 7256–7262.Google Scholar
  15. 15.
    Sanchez-Murcia, P. A., Bueren-Calabuig, J. A., Camacho-Artacho, M., Cortes-Cabrera, A., & Gago, F. (2016). Stepwise simulation of 3,5-dihydro-5-methylidene-4H-imidazol-4-one (MIO) biogenesis in histidine ammonia-lyase. Biochemistry, 55(41), 5854–5864.Google Scholar
  16. 16.
    Watts, K. T., Mijts, B. N., Lee, P. C., Manning, A. J., & Schmidt-Dannert, C. (2006). Discovery of a substrate selectivity switch in tyrosine ammonia-lyase, a member of the aromatic amino acid lyase family. Chemistry & Biology, 13(12), 1317–1326.Google Scholar
  17. 17.
    Heberling, M. M., Masman, M. F., Bartsch, S., Wybenga, G. G., Dijkstra, B. W., Marrink, S. J., & Janssen, D. B. (2015). Ironing out their differences: dissecting the structural determinants of a phenylalanine aminomutase and ammonia lyase. ACS Chemical Biology, 10(4), 989–997.Google Scholar
  18. 18.
    Walter, T., King, Z., & Walker, K. D. (2016). A tyrosine aminomutase from rice (Oryza sativa) isomerizes (S)-alpha- to (R)-beta-tyrosine with unique high enantioselectivity and retention of configuration. Biochemistry, 55(1), 1–4.Google Scholar
  19. 19.
    Lovelock, S. L., Lloyd, R. C., & Turner, N. J. (2014). Phenylalanine ammonia lyase catalyzed synthesis of amino acids by an MIO-cofactor independent pathway. Angewandte Chemie, International Edition, 53(18), 4652–4656.Google Scholar
  20. 20.
    Pinto, G. P., Ribeiro, A. J. M., Ramos, M. J., Fernandes, P. A., Toscano, M., & Russo, N. (2015). New insights in the catalytic mechanism of tyrosine ammonia-lyase given by QM/MM and QM cluster models. Archives of Biochemistry and Biophysics, 582, 107–115.Google Scholar
  21. 21.
    Bencze, L. C., Filip, A., Banoczi, G., Tosa, M. I., Irimie, F. D., Gellert, A., Poppe, L., & Paizs, C. (2017). Expanding the substrate scope of phenylalanine ammonia-lyase from Petroselinum crispum towards styrylalanines. Organic & Biomolecular Chemistry, 15(17), 3717–3727.Google Scholar
  22. 22.
    Jaliani, H. Z., Farajnia, S., Mohammadi, S. A., Barzegar, A., & Talebi, S. (2013). Engineering and kinetic stabilization of the therapeutic enzyme Anabeana variabilis phenylalanine ammonia lyase. Applied Biochemistry and Biotechnology, 171(7), 1805–1818.Google Scholar
  23. 23.
    Lovelock, S. L., & Turner, N. J. (2014). Bacterial Anabaena variabilis phenylalanine ammonia lyase: A biocatalyst with broad substrate specificity. Bioorganic and Medicinal Chemistry, 22(20), 5555–5557.Google Scholar
  24. 24.
    DreSsen, A., Hilberath, T., Mackfeld, U., Billmeier, A., Rudat, J., & Pohl, M. (2017). Phenylalanine ammonia lyase from Arabidopsis thaliana (AtPAL2): a potent MIO-enzyme for the synthesis of non-canonical aromatic alpha-amino acids. Part I: Comparative characterization to the enzymes from Petroselinum crispum (PcPAL1) and Rhodosporidium toruloides (RtPAL). Journal of Biotechnology, 258, 148–157.Google Scholar
  25. 25.
    DreSsen, A., Hilberath, T., Mackfeld, U., Billmeier, A., Rudat, J., & Pohl, M. (2017). Phenylalanine ammonia lyase from Arabidopsis thaliana (AtPAL2): a potent MIO-enzyme for the synthesis of non-canonical aromatic alpha-amino acids. Part II: Application in different reactor concepts for the production of (S)-2-chloro-phenylalanine. Journal of Biotechnology, 258, 158–166.Google Scholar
  26. 26.
    Parmeggiani, F., Lovelock, S. L., Weise, N. J., Ahmed, S. T., & Turner, N. J. (2015). Synthesis of D- and L-phenylalanine derivatives by phenylalanine ammonia lyases: a multienzymatic cascade process. Angewandte Chemie, International Edition, 54(15), 4608–4611.Google Scholar
  27. 27.
    Liu, L., Hossain, G. S., Shin, H. D., Li, J. H., Du, G. C., & Chen, J. (2013). One-step production of alpha-ketoglutaric acid from glutamic acid with an engineered L-amino acid deaminase from Proteus mirabilis. Journal of Biotechnology, 164(1), 97–104.Google Scholar
  28. 28.
    Motta, P., Molla, G., Pollegioni, L., & Nardini, M. (2016). Structure-function relationships in L-amino acid deaminase, a flavor protein belonging to a novel class of biotechnologically relevant enzymes. The Journal of Biological Chemistry, 291(20), 10457–10475.Google Scholar
  29. 29.
    Baek, J. O., Seo, J. W., Kwon, O., Seong, S. I., Kim, I. H., & Kim, C. H. (2011). Expression and characterization of a second L-amino acid deaminase isolated from Proteus mirabilis in Escherichia coli. Journal of Basic Microbiology, 51(2), 129–135.Google Scholar
  30. 30.
    Fukuhara, T. Y. S. (1990). Novel ligand-exchange chromatographic resolution of DL-amino acids using nucleotides and coenzymes. Journal of Chromatographic Science, 28(1), 114–117.Google Scholar
  31. 31.
    Wybenga, G. G., Szymanski, W., Wu, B., Feringa, B. L., Janssen, D. B., & Dijkstra, B. W. (2014). Structural investigations into the stereochemistry and activity of a phenylalanine-2,3-aminomutase from Taxus chinensis. Biochemistry, 53(19), 3187–3198.Google Scholar
  32. 32.
    Ratnayake, N. D., Wanninayake, U., Geiger, J. H., & Walker, K. D. (2011). Stereochemistry and mechanism of a microbial phenylalanine aminomutase. Journal of the American Chemical Society, 133(22), 8531–8533.Google Scholar
  33. 33.
    Mutatu, W., Klettke, K. L., Foster, C., & Walker, K. D. (2007). Unusual mechanism for an aminomutase rearrangement: retention of configuration at the migration termini. Biochemistry, 46(34), 9785–9794.Google Scholar
  34. 34.
    Feng, L., Wanninayake, U., Strom, S., Geiger, J., & Walker, K. D. (2011). Mechanistic, mutational, and structural evaluation of a Taxus phenylalanine aminomutase. Biochemistry, 50(14), 2919–2930.Google Scholar
  35. 35.
    Alexandre, F. R., Pantaleone, D. P., Taylor, P. P., Fotheringham, I. G., Ager, D. J., & Turner, N. J. (2002). Amine-boranes: effective reducing agents for the deracemisation of DL-amino acids using L-amino acid oxidase from Proteus myxofaciens. Tetrahedron Letters, 43(4), 707–710.Google Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  • Longbao Zhu
    • 1
  • Guoqiang Feng
    • 1
  • Fei Ge
    • 1
  • Ping Song
    • 1
  • Taotao Wang
    • 1
  • Yi Liu
    • 2
    Email author
  • Yugui Tao
    • 1
  • Zhemin Zhou
    • 3
  1. 1.School of Biochemical EngineeringAnhui Polytechnic UniversityWuhuPeople’s Republic of China
  2. 2.Key Laboratory of Food and Biotechnology, School of Food and BiotechnologyXihua UniversityChengduChina
  3. 3.Key Laboratory of Industrial Biotechnology, Ministry of Education, School of BiotechnologyJiangnan UniversityWuxiPeople’s Republic of China

Personalised recommendations