Enantioselective synthesis of vicinal R,R-diols by yeast butanediol dehydrogenaseReport as inadecuate




Enantioselective synthesis of vicinal R,R-diols by yeast butanediol dehydrogenase - Download this document for free, or read online. Document in PDF available to download.

* Corresponding author 1 Department of Biochemistry and Molecular Biology 2 SPO - Sciences Pour l-Oenologie 3 Department of Chemistry

Abstract : 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 NADH 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.





Author: Eduard Calam - Eva González-Roca - M Rosario Fernández - Sylvie Dequin - Xavier Parés - Albert Virgili - Josep A Biosca -

Source: https://hal.archives-ouvertes.fr/



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