Invited Talk 21st International Conference on Biological Inorganic Chemistry 2025

Elucidating Reaction Mechanisms of Coupled Binuclear Copper Enzymes by Correlating QM/MM Calculations, Spectroscopy, and Kinetics (123964)

Lubomír Rulíšek 1 , Agnieszka Stańczak 1 , Ioannis Kipouros 2 , Edward I Solomon 3
  1. Institute of Organic Chemistry and Biochemistry, Czech Academy of Sciences, Praha 6, Czech Republic
  2. Dept of Chemistry, 385 Frick Laboratory, Princeton University, Princeton, New Jersey, United States of America
  3. Department of Chemistry, Stanford University, Stanford, California, United States of America

Coupled binuclear copper (CBC) enzymes are used by nature to catalyze variety of chemical conversions (e.g., hydroxylations, oxidations).[1] One such example is tyrosinase (Ty), the ubiquitous enzyme responsible for O2-dependent ortho-hydroxylation of L-tyrosine to L-3,4-dihydroxyphenylalanine (L-DOPA), as well as the subsequent two-electron oxidation of L-DOPA to L-dopaquinone. These elementary reactions are the initial and rate-limiting steps in melanogenesis.[1] Due to its role in the biosynthesis of melanin, Ty is of an interest in the detection, prevention, and treatment of complex human diseases, including skin cancer [2] and Parkinson’s disease.[3] By combining theoretical and experimental methods we investigated H-bonding interactions in oxy-Ty [Ty/O2] active site, [4] and predicted structure of the elusive ternary complex [Ty/O2/monophenol] intermediate.[5] Next, we examined monooxygenation reaction of Ty with the analogue of native substrate, that is with methyl 4-hydroxybenzoate.[5] Subsequently, we investigated the ortho-hydroxylation step employing series of substrates with different electron donating/withdrawing group in phenol para-position revealing biphasic substrate dependence of the monophenol monooxygenation reaction of tyrosinase. This biphasic nature is ascertained mainly by correlating experimental and theoretical energy barrier/k2, and solvent KIE. [6] Currently, concerted compuational and experimental (XAS and EPR spectroscopy, and kinetics) are underway [7] to provide the comprehensive reaction cycle of Ty that may open an avenue to structure/function correlations across the family of CBC enzymes.[7,8]

 

[1] E. I. Solomon et al., Chem. Rev. 2014, 114, 3659–3853.

[2] B. Ciui et al., Adv. Healthcare Mater. 2018, 7, 1701264.

[3] I. Carballo-Carbajal et al., Nat. Commun. 2019, 10, 973.

[4] I. Kipouros, A. Stańczak, L. Rulíšek, E. I. Solomon et al., Chem. Commun. 2022, 58, 3913–3916.

[5] I. Kipouros, A. Stańczak, L. Rulíšek, E. I. Solomon et al., Proc. Natl. Acad. Sci. U. S. A. 2022, 119, e2205619119.

[6] I. Kipouros, A. Stańczak, L. Rulíšek, E. I. Solomon et al. J. Am. Chem. Soc. 2023, 145, 22866–22870.

[7] A. Stańczak, I. Kipouros, P. Eminger, E. M. Dunietz, E. I. Solomon, L. Rulíšek Coord. Chem. Rev. 2025, 525, 216301

[8] A. Stańczak, I. Kipouros, L. D. Elmendorf, E. I. Solomon, , L. Rulíšek et al., to be submitted