Enantioselective synthesis of vicinal (R,R)-diols by yeast butanediol dehydrogenase

Autor: Calam, Eduard, González-Roca, Eva, Fernández, M Rosario, Dequin, Sylvie, Parés, Xavier, Virgili, Albert, Biosca, Josep A
Přispěvatelé: Department of Biochemistry and Molecular Biology, Universitat Autònoma de Barcelona [Barcelona] (UAB), Sciences Pour l'Oenologie (SPO), Institut National de la Recherche Agronomique (INRA)-Université de Montpellier (UM)-Université Montpellier 1 (UM1)-Institut de Recherche pour le Développement (IRD [Nouvelle-Calédonie])-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), Department of Chemistry, Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro), Ministry of Education and Science of Spain [BIO2007-64659 ], Generalitat de Catalunya (Government of Catalonia) [2009 SGR795], Universitat Autònoma de Barcelona (UAB), Université Montpellier 1 (UM1)-Institut National de la Recherche Agronomique (INRA)-Université de Montpellier (UM)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro)
Jazyk: angličtina
Rok vydání: 2016
Předmět:
Zdroj: Applied and Environmental Microbiology
Applied and Environmental Microbiology, American Society for Microbiology, 2016, ⟨10.1128/AEM.03717-15⟩
Applied and Environmental Microbiology, American Society for Microbiology, 2016, 82 (6), ⟨10.1128/AEM.03717-15⟩
ISSN: 0099-2240
1098-5336
DOI: 10.1128/AEM.03717-15⟩
Popis: Butanediol dehydrogenase (Bdh1p) from Saccharomyces cerevisiae belongs to the superfamily of the medium chain dehydrogenases/reductases and converts reversibly R-acetoin and S-acetoin to (2R,3R)-2,3-butanediol and meso-2,3-butanediol, respectively. It is specific for NAD(H) as a coenzyme and it is the main enzyme involved in the last metabolic step leading to (2R,3R)-2,3-butanediol in yeast. In this study, we have used the activity of Bdh1p, in different forms: purified enzyme, yeast extracts, permeabilized yeast cells, and as a fusion protein (with yeast formate dehydrogenase, Fdh1p) to transform several vicinal diketones to the corresponding diols. We have also developed a new variant of the "delitto perfetto" methodology to place BDH1 under the control of the GAL1 promoter, resulting in a yeast strain that overexpresses butanediol dehydrogenase and formate dehydrogenase activities in the presence of galactose and regenerates NADH in the presence of formate. While the use of purified Bdh1p allows the synthesis of enantiopure (2R,3R)-2,3-butanediol, (2R,3R)-2,3-pentanediol, (2R,3R)-2,3-hexanediol and (3R,4R)-3,4-hexanediol, the use of the engineered strain (as an extract or as permeabilized cells), yields mixtures of the diols. The production of pure diol stereoisomers has also been achieved by means of a chimeric fusion protein combining Fdh1p and Bdh1p. Finally, we have determined the selectivity of Bdh1p towards the oxidation/reduction of the hydroxyl/ketone groups from (2R,3R)-2,3-pentanediol/2,3-pentanedione and (2R,3R)-2,3-hexanediol/2,3-hexanedione. In conclusion, Bdh1p is an enzyme with biotechnological interest that can be used to synthesize chiral building blocks. A scheme of the favored pathway with the corresponding intermediates is proposed for the Bdh1p reaction.
Databáze: OpenAIRE
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