39516-03-5Relevant articles and documents
Discovery and Redesign of a Family VIII Carboxylesterase with High (S)-Selectivity toward Chiral sec-Alcohols
Park, Areum,Park, Seongsoon
, p. 2397 - 2402 (2022/02/17)
Highly enantioselective lipase has been widely utilized in the preparation of versatile enantiopure chiral sec-alcohols through kinetic or dynamic kinetic resolution. Lipase is intrinsically (R)-selective, and it is difficult to obtain (S)-selective lipase. Recent crystal structures of a family VIII carboxylesterase have revealed that the spatial array of its catalytic triad is the mirror image of that of lipase but with a catalytic triad that is distinct from lipase. We, therefore, hypothesized that the family VIII carboxylesterase may exhibit (S)-enantioselectivity toward sec-alcohols similar to (S)-selective serine protease, whose catalytic triad is also spatially arrayed as its mirror image. In this study, a homologous enzyme (carboxylesterase from Proteobacteria bacterium SG_bin9, PBE) of a known family VIII carboxylesterase (pdb code: 4IVK) was prepared, which showed not only moderate (S)-selectivity toward sec-alcohols such as 3-butyn-2-ol and 1-phenylethyl alcohol but also (R)-selectivity toward particular sec-alcohols among the substrates explored. Furthermore, the (S)-selectivity of PBE has been significantly improved by rational redesign based on molecular modeling. Molecular modeling identified a binding pocket composed of Ser381, Ala383, and Arg408 for the methyl substituent of (R)-1-phenylethyl acetate and suggested that larger residues may increase the enantioselectivity by interfering with the binding of the slow-reacting enantiomer. As predicted, substituting Ser381with larger residues (Phe, Tyr, and Trp) significantly improved the (S)-selectivity of PBE toward all sec-alcohols explored, even the substrates toward which the wild-type PBE exhibits (R)-selectivity. For instance, the enantioselectivity toward 3-butyn-2-ol and 1-phenylethyl alcohol was improved from E = 5.5 and 36.1 to E = 2001 and 882, respectively, by single mutagenesis (S381F).
Application of robust ketoreductase from Hansenula polymorpha for the reduction of carbonyl compounds
Petrovi?ová, Tatiana,Gyuranová, Dominika,Pl?, Michal,Myrtollari, Kamela,Smonou, Ioulia,Rebro?, Martin
, (2021/02/05)
Enzyme-catalysed asymmetric reduction of ketones is an attractive tool for the production of chiral building blocks or precursors for the synthesis of bioactive compounds. Expression of robust ketoreductase (KRED) from Hansenula polymorpha was upscaled and applied for the asymmetric reduction of 31 prochiral carbonyl compounds (aliphatic and aromatic ketones, diketones and β-keto esters) to the corresponding optically pure hydroxy compounds. Biotransformations were performed with the purified recombinant KRED together with NADP+ recycling glucose dehydrogenase (GDH, Bacillus megaterium), both overexpressed in Escherichia coli BL21(DE3). Maximum activity of KRED for biotransformation of ethyl-2-methylacetoacetate achieved by the high cell density cultivation was 2499.7 ± 234 U g–1DCW and 8.47 ± 0.40 U·mg–1E, respectively. The KRED from Hansenula polymorpha is a very versatile enzyme with broad substrate specificity and high activity towards carbonyl substrates with various structural features. Among the 36 carbonyl substrates screened in this study, the KRED showed activity with 31, with high enantioselectivity in most cases. With several ketones, the Hansenula polymorpha KRED catalysed preferentially the formation of the (R)-secondary alcohols, which is highly valued.
Impact of the Difluoromethylene Group in the Organocatalyzed Acylative Kinetic Resolution of α,α-Difluorohydrins
Andrei, Daniela,Bressy, Cyril,Desrues, Titouan,Médebielle, Maurice,Merad, Jérémy,Parrain, Jean-Luc,Pons, Jean-Marc,Quintard, Adrien
supporting information, p. 24924 - 24929 (2021/10/19)
Due to the omnipresence of chiral organofluorine compounds in pharmaceutical, agrochemical, and material chemistry, the development of enantioselective methods for their preparation is highly desirable. In the present study, the enantioselective organocatalyzed acylation of α,α-difluorohydrins using a commercially available chiral isothiourea is reported through a kinetic resolution (KR) process. It reveals that the difluoromethylene moiety (C(sp3)F2) can serve as a directing group through electrostatic fluorine–cation interactions, greatly improving the enantioselectivity of the KR. In this context, a broad range of fluorinated alcohols such as valuable 4,4-difluoro-1,3-diols could be synthesized with exquisite enantiocontrol (typically >99:1 er). Turning to 2,2-difluoro-1,3-diols, we also demonstrated that aromatic and fluorinated groups were mutually compatible to provide the expected enantioenriched adducts with >99:1 er.
