Deuterium isotope effects in mechanistic studies of biotransformations of l-tyrosine and p-hydroxyphenylpyruvic acid catalyzed by the enzyme l-phenylalanine dehydrogenase

Katarzyna Pałka; Katarzyna Podsadni; Jolanta Szymańska-Majchrzak; Elżbieta Winnicka

doi:10.2478/nuka-2025-0006

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Deuterium isotope effects in mechanistic studies of biotransformations of l-tyrosine and p-hydroxyphenylpyruvic acid catalyzed by the enzyme l-phenylalanine dehydrogenase

Nukleonika

Volume 70 (2025): Issue 2 (June 2025)

By: Katarzyna Pałka , Katarzyna Podsadni, Jolanta Szymańska-Majchrzak and Elżbieta Winnicka

Open Access

|May 2025

Abstract

The mechanisms of the reversible oxidative deamination of l-tyrosine to p-hydroxyphenylpyruvic acid and reductive amination of phenylpyruvic acid to l-phenylalanine, both catalyzed by the enzyme l-phenylalanine dehydrogenase (PheDH, EC 1.4.1.20), were investigated using the kinetic isotope effect (KIE) and solvent isotope effect (SIE) methods. The values of deuterium kinetic effects in the 2-position of l-tyrosine and KIE in the (3S)-position of phenylpyruvic acid and solvent isotope effects for both reactions were determined using the non-competitive spectrophotometric method. Some mechanistic details of these biotransformations were discussed.

References

Scriver, C. R. (2007). The PAH gene, phenylketonuria, and a paradigm shift. Hum. Mutat., 28(9), 831–845.
Search in Google Scholar Back to article
Williams, R. A., Mamotte, C. D. S., & Burnett, J. R. (2008). Phenylketonuria: An inborn error of phenylalanine metabolism. Clin. Biochem. Rev., 29(1), 31–41.
Search in Google Scholar Back to article
Hendriksz, C. J., & Walter, J. H. (2004). Update on phenylketonuria. Curr. Pediatr., 14(5), 400–406.
Search in Google Scholar Back to article
Mitchell, G. A., Grompe, M., Lambert, M., & Tanguay, R. M. (2001) Hypertyrosinemia. In C. R. Scriver, A. L. Beaudet & W. S. Sly (Eds.), The metabolic and molecular bases of inherited disease. (8th ed., Vol. II, pp. 1777–1785). New York: McGraw-Hill.
Search in Google Scholar Back to article
Brunhuber, N. M. W., Banerjee, A., Jacobs, W. R. Jr, & Blanchard, J. S. (1994). Cloning, sequencing, and expressing of Rhodococcus l-phenylalanine dehydrogenase. J. Biol. Chem., 269(23), 16203–16211.
Search in Google Scholar Back to article
Brunhuber, N. M. W., Thoden, J. B., Blanchard, J. S., & Vanhooke, J. L. (2000). Rhodococcus l-phenylalanine dehydrogenase: kinetics, mechanism, and structural basis for catalytic specifity. Biochemistry, 39(31), 9174–9187.
Search in Google Scholar Back to article
Seah, S. Y. K., Britton, K. L., Rice, D. W., Asano, Y., & Engel, P. C. (2002). Single amino acid substitution in Bacillus sphaericus phenylalanine dehydrogenase dramatically increases its discrimination between phenylalanine and tyrosine substrates. Biochemistry, 41, 11390–11397.
Search in Google Scholar Back to article
Seah, S. Y. K., Britton, K. L., Rice, D. W., Asano, Y., & Engel, P. C. (2003). Kinetic analysis of phenylalanine dehydrogenase mutants designed for aliphatic amino acid dehydrogenase with guidance from homologybased modelling. Eur. J. Biochem., 270, 4628–4634.
Search in Google Scholar Back to article
Asano, Y., Yamada, A., Kato, Y., Yamaguchi, K., Hibino, Y., Hirai, K., & Kondo, K. (1990). Enantioselective synthesis of (S)-amino acids by phenylalanine dehydrogenase from Bacillus sphaericus: use of natural and recombinant enzymes. J. Org. Chem., 55(21), 5567–5571.
Search in Google Scholar Back to article
Busca, P., Paradisi, F., Moynihan, E., Maguire, A. R., & Engel, P. C. (2004). Enantioselective synthesis of non-natural amino acids using phenylalanine dehydrogenase modified by site-directed mutagenesis. Org. Biomol. Chem., 2, 2684–2691.
Search in Google Scholar Back to article
Hummel, W. E., Schmidt, E., Wandrey, C., & Kula, M.-R. (1986). l-Phenylalanine dehydrogenase from Brevibacterium sp. for production l-phenylalanine by reductive amination of phenylpyruvate. Appl. Microbiol. Biotechnol., 25(3), 175–185.
Search in Google Scholar Back to article
Sühnel, J. R. L., & Schowen, L. R. (1991). Theoretical basis for primary and secondary hydrogen isotope effects. In P. F. Cook (Ed.), Enzyme mechanism from isotope effects (pp. 3–35). Boca Raton (FL): CRC Press.
Search in Google Scholar Back to article
Schowen, L. R. (1972). Mechanistic deductions from solvent isotope effect. Prog. Phys. Org. Chem., 9, 275–332.
Search in Google Scholar Back to article
Jemielity, J., Kański, R., & Kańska, M. (2001). Synthesis of tritium labeled [3R-₃H]-, and [3S-₃H]-l-phenylalanine. J. Label. Compd. Radiopharm., 44, 205–304.
Search in Google Scholar Back to article
Skowera, K., & Kańska, M. (2008). Enzymatic synthesis of phenylpyruvic acid labeled with deuterium, tritium, and carbon-14. J. Label. Compd., 51, 321–324.
Search in Google Scholar Back to article
Pałka, K., & Kańska, M. (2012). Enzymatic reductive amination of p-hydroxy- and phenylpyruvic acids as methods of synthesis of l-tyrosine and l-phenylalanine labeled with deuterium and tritium. Nukleonika, 57(3), 383–387.
Search in Google Scholar Back to article
Gary, R., Bates, R. G., & Robinson, R. A. (1964). Second dissociation constant of deuteriophosphoric acid in deuterium oxide from 5 to 50°C: Standardization of pD scale. J. Phys. Chem., 68(12), 3806–3809.
Search in Google Scholar Back to article
Kańska, M., Dragulska, S., Pająk, M., & Winnicka, E. (2015). Isotope effects in the hydroxylation of ltyrosine catalyzed by tyrosinase. J. Radioanal. Nucl. Chem., 305(2), 371–378.
Search in Google Scholar Back to article
Parkin, D. W. (1991). Methods for determination of competitive and noncompetitive kinetics isotope effects. In P. F. Cook (Ed.), Enzyme mechanism from isotope effects (pp. 269–290), Boca Raton (FL): CRC Press.
Search in Google Scholar Back to article
Papajak, E., Kwiecień, R. A., Rudziński, J., Sicińska, D., Kamiński, R., Szadkowski, Ł., Kurihara, T., Esaki, N., & Paneth, P. (2006). Mechanism of reaction catalyzed by DL-2-haloacid dehalogenase from kinetic isotope effects. Biochemistry, 45(19), 6012–6017.
Search in Google Scholar Back to article
Brunhuber, N. M. W., & Blanchard, J. S. (1994). The biochemistry and enzymology of amino acid dehydrogenases. Crit. Rev. Biochem. Mol. Biol., 29(6), 415–467.
Search in Google Scholar Back to article
Wende, U., Koppelkam, M., Hummel, W., Sander, J., & Langenbeck, U. (1990). A new approach to the newborn screening for hyperphenylalaninemias: use of l-phenylalanine dehydrogenase and micrititer plates. Clin. Chim. Acta, 192(3), 165–170.
Search in Google Scholar Back to article
Naghib, S. M., Rabee, M., Omidinia, E., & Khoshkenar, P. (2012). Investigation of a biosensor based on phenylalanine dehydrogenase immobilized on the polymer-blend film for phenylketonuria. Electroanalysis, 24, 407–417.
Search in Google Scholar Back to article
Asano, Y., & Nakazawa, A. (1987). High yield synthesis of l-amino acids by phenylalanine dehydrogenase from Sporasacrina ureae. Agric. Biol. Chem., 51(7), 2035–2036.
Search in Google Scholar Back to article
Vanhooke, J. L., Thoden, J. B., Brunhuber, N. M. W., Blanchard, J. S., & Holden, H. M. (1999). Phenylalanine dehydrogenase from Rhodococcus sp. M4: High-resolution X-ray analyses of inhibitory ternary complexes reveal key features in the oxidative deamination mechanism. Biochemistry, 38(8), 2326–2339.
Search in Google Scholar Back to article

Articles in this issue

DOI: https://doi.org/10.2478/nuka-2025-0006 | Journal eISSN: 1508-5791 | Journal ISSN: 0029-5922

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Language: English

Page range: 51 - 56

Submitted on: Jan 5, 2024

Accepted on: Mar 4, 2025

Published on: May 2, 2025

Published by: Institute of Nuclear Chemistry and Technology

In partnership with: Paradigm Publishing Services

Publication frequency: 4 issues per year

Keywords:

Kinetic isotope effects,

l-Phenylalanine,

Phenylalanine dehydrogenase,

p-Hydroxyphenylpyruvic acid,

Solvent isotope effects,

Tyrosine

Related subjects:

Chemistry,

Nuclear chemistry,

Physics,

Astronomy and astrophysics,

Physics, other

© 2025 Katarzyna Pałka, Katarzyna Podsadni, Jolanta Szymańska-Majchrzak, Elżbieta Winnicka, published by Institute of Nuclear Chemistry and Technology
This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 License.

Volume 70 (2025): Issue 2 (June 2025)