Structural analysis of an anthrol reductase inspires enantioselective synthesis of enantiopure hydroxycycloketones and β-halohydrins

Abstract

Asymmetric reduction of prochiral ketones, particularly, reductive desymmetrization of 2,2-disubstituted prochiral 1,3-cyclodiketones to produce enantiopure chiral alcohols is challenging. Herein, an anthrol reductase CbAR with the ability to accommodate diverse bulky substrates, like emodin, for asymmetric reduction is identified. We firstly solve crystal structures of CbAR and CbAR-Emodin complex. It reveals that Tyr210 is critical for emodin recognition and binding, as it forms a hydrogen-bond interaction with His162 and π-π stacking interactions with emodin. This ensures the correct orientation for the stereoselectivity. Then, through structure-guided engineering, variant CbAR-H162F can convert various 2,2-disubstituted 1,3-cyclodiketones and α-haloacetophenones to optically pure (2S, 3S)-ketols and (R)-β-halohydrins, respectively. More importantly, their stereoselectivity mechanisms are also well explained by the respective crystal structures of CbAR-H162F-substrate complex. Therefore, this study demonstrates that an in-depth understanding of catalytic mechanism is valuable for exploiting the promiscuity of anthrol reductases to prepare diverse enantiopure chiral alcohols.

Anthrol reductases: discovery, role in biosynthesis and applications in natural product syntheses

“…JNU001 (CbAR), which is supposed to be involved in the biosynthesis of phytotoxin beticolin 1. 61,62 CbAR shows high sequence similarity with known anthrol reductases and 79.2% and 83.9% sequence similarity with ARti-2 and ARti from T. islandicus. 62 CbAR has been shown to efficiently catalyze the reduction of emodin hydroquinone to 14a with >99% conversion and >99% ee under optimal condition (pH 8.0, 25 °C) using NADPH in the presence of Na 2 S 2 O 4 (Scheme 13).…”

“…61,62 CbAR shows high sequence similarity with known anthrol reductases and 79.2% and 83.9% sequence similarity with ARti-2 and ARti from T. islandicus. 62 CbAR has been shown to efficiently catalyze the reduction of emodin hydroquinone to 14a with >99% conversion and >99% ee under optimal condition (pH 8.0, 25 °C) using NADPH in the presence of Na 2 S 2 O 4 (Scheme 13). 62 To explain the catalytic mechanism of CbAR, the crystal structure for CbAR-NADP + and CbAR-NADP +emodin complexes were determined, with 3.30 Å and 1.85 Å resolutions, respectively.…”

“…Therefore, the mutation of either His162 or Tyr210 results in a complete loss of the reduction activity of CbAR. 62 The residue Asn152 also binds with the emodin through H-bonded interaction (Fig. 4B).…”

Semirational Engineering of a Thermostable Carbonyl Reductase for the Precision Synthesis of (2R,3R)-2-Methyl-2-benzyl-3-hydroxycyclopentanone and Its Analogues

“…(2 R ,3 R )-Ketol is the key chiral intermediate for the synthesis of (+)-estrone and cortistatins. Single stereoisomers of these hydroxyketones are highly demanded, and desymmetric reduction of prochiral 2,2-disubstituted-1,3-cyclopentanediones is a direct and powerful approach to access the single stereoisomers of 2,2-disubstituted-3-hydroxycyclopentanones. − For chemical asymmetric reduction, (2 S ,3 S )-2-methyl-2-benzyl-3-hydroxycyclopentanone ( 2a ) was obtained in 96% ee and 49:1 dr when chiral phosphiamid was employed as a catalyst . Using the Ir/f-ampha complex or DIBAL-H as the catalyst, (2 R ,3 R )- 2a was obtained with 99% ee and 21:1 dr or 74% ee and 4:1 dr, respectively (Figure B).…”

“…16 Using the Ir/ f-ampha complex 12 or DIBAL-H 17 as the catalyst, (2R,3R)-2a was obtained with 99% ee and 21:1 dr or 74% ee and 4:1 dr, respectively (Figure 1B). For the biocatalytic reduction, through the structure-guided directed evolution of carbonyl reductases RasADH, 13 SSCR, 14 and CbAR, 15 the highly efficient and stereoselective reduction of 2,2-substituted-1,3cyclopentanediones was realized to give the corresponding (2R,3S)-or (2S,3S)-ketols as a single stereoisomer in many cases (Figure 1B). Herein, we report the biocatalytic desymmetric reduction of 2,2-disubstituted-1,3-cyclopentanediones to afford the (2R,3R)-stereoisomers of 2,2-disubstituted-3-hydroxycyclopentanones by semirational engineering of the carbonyl reductase CBCR (WP_026035294.1) from Cupriavidus sp.…”