Enantiomerically enriched hydroxyketones are well-established intermediates for the synthesis of several bioactive compounds [1] and can be chemically obtained by stereoselective reduction of one of the carbonyl moieties of the corresponding diketones. However, enzymatic strategies are characterized by higher catalytic efficiency, milder reaction conditions, higher stereo- and regioselectivity, and fewer numbers of synthetic steps. Therefore, they can be chosen as convenient and environmentally friendly alternatives.[2] A NADPH-dependent benzil reductase from the non-conventional yeast Pichia glucozyma (KRED1-Pglu) was over-expressed in E. coli, purified and exploited to catalyze the asymmetric monoreduction of bulky aromatic 1,2-dicarbonyl compounds. The cofactor was recycled by an enzyme-coupled system (glucose-glucose dehydrogenase (GDH) from Bacillus megaterium). The recombinant KRED1-Pglu showed a wide range of activity (24-97% conversion) and excellent stereoselectivity (ee ≥ 96% in all but one case). On the contrary, it proved either inactive or very poorly active towards most 1,3- and 1,4-dicarbonyls tested as potential substrates. In order to understand this peculiar behavior, the enzyme was crystallized (1.77 Å resolution) and its active site was investigated to identify the recognition residues involved in the desymmetrization reaction. QM and classical calculations also allowed for a proposal of the catalytic mechanism, along with an in silico reactivity prediction.[3] [1] G. Aullón, P. Romea, F. Urpí, Synthesis 2017, 49, 484. [2] P. Hoyos, J.-V. Sinisterra, F. Molinari, A. R. Alántara, P. Domínguez de María, Acc. Chem. Res. 2010, 43, 288. [3] M. Rabuffetti, P. Cannazza, M. L. Contente, A. Pinto, D. Romano, P. Hoyos, A. R. Alcántara, I. Eberini, T. Laurenzi, L. Gourlay, F. Di Pisa, F. Molinari, Bioorg. Chem. 2021, 108, 104644.

Stereoselective monoreduction of bulky 1,2-dicarbonyls catalyzed by a benzil reductase from Pichia glucozyma (KRED1-Pglu) / M. Rabuffetti, P. Cannazza, M.L. Contente, A. Pinto, D. Romano, P. Hoyos, A.R. Alcántara, I. Eberini, T. Laurenzi, L.J. Gourlay, F. DI PISA, F.E. Molinari. ((Intervento presentato al 27. convegno Congresso Nazionale della Società Chimica Italiana-SCI tenutosi a online nel 2021.

Stereoselective monoreduction of bulky 1,2-dicarbonyls catalyzed by a benzil reductase from Pichia glucozyma (KRED1-Pglu)

M. Rabuffetti;P. Cannazza;M.L. Contente;A. Pinto;D. Romano;I. Eberini;T. Laurenzi;L.J. Gourlay;F. DI PISA;F.E. Molinari
2021

Abstract

Enantiomerically enriched hydroxyketones are well-established intermediates for the synthesis of several bioactive compounds [1] and can be chemically obtained by stereoselective reduction of one of the carbonyl moieties of the corresponding diketones. However, enzymatic strategies are characterized by higher catalytic efficiency, milder reaction conditions, higher stereo- and regioselectivity, and fewer numbers of synthetic steps. Therefore, they can be chosen as convenient and environmentally friendly alternatives.[2] A NADPH-dependent benzil reductase from the non-conventional yeast Pichia glucozyma (KRED1-Pglu) was over-expressed in E. coli, purified and exploited to catalyze the asymmetric monoreduction of bulky aromatic 1,2-dicarbonyl compounds. The cofactor was recycled by an enzyme-coupled system (glucose-glucose dehydrogenase (GDH) from Bacillus megaterium). The recombinant KRED1-Pglu showed a wide range of activity (24-97% conversion) and excellent stereoselectivity (ee ≥ 96% in all but one case). On the contrary, it proved either inactive or very poorly active towards most 1,3- and 1,4-dicarbonyls tested as potential substrates. In order to understand this peculiar behavior, the enzyme was crystallized (1.77 Å resolution) and its active site was investigated to identify the recognition residues involved in the desymmetrization reaction. QM and classical calculations also allowed for a proposal of the catalytic mechanism, along with an in silico reactivity prediction.[3] [1] G. Aullón, P. Romea, F. Urpí, Synthesis 2017, 49, 484. [2] P. Hoyos, J.-V. Sinisterra, F. Molinari, A. R. Alántara, P. Domínguez de María, Acc. Chem. Res. 2010, 43, 288. [3] M. Rabuffetti, P. Cannazza, M. L. Contente, A. Pinto, D. Romano, P. Hoyos, A. R. Alcántara, I. Eberini, T. Laurenzi, L. Gourlay, F. Di Pisa, F. Molinari, Bioorg. Chem. 2021, 108, 104644.
Settore CHIM/06 - Chimica Organica
Settore CHIM/11 - Chimica e Biotecnologia delle Fermentazioni
Società Chimica Italiana
http://www.sci2020.org/
Stereoselective monoreduction of bulky 1,2-dicarbonyls catalyzed by a benzil reductase from Pichia glucozyma (KRED1-Pglu) / M. Rabuffetti, P. Cannazza, M.L. Contente, A. Pinto, D. Romano, P. Hoyos, A.R. Alcántara, I. Eberini, T. Laurenzi, L.J. Gourlay, F. DI PISA, F.E. Molinari. ((Intervento presentato al 27. convegno Congresso Nazionale della Società Chimica Italiana-SCI tenutosi a online nel 2021.
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/2434/877018
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