Discovery of a novel ortho-haloacetophenones-specific carbonyl reductase from Bacillus aryabhattai and insight into the molecular basis for its catalytic performance

Li, Aipeng; Yuchi, Qingxiao; Li, Xue; Pang, Wei; Li, Bin; Xue, Feng; Zhang, Lianbing

doi:10.1016/j.ijbiomac.2019.07.153

Cited by 13 publications

(15 citation statements)

References 57 publications

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“…This provided ag ood case to investigate the relationship between substrate structure and enzymes tereoselectivity.T oc larify the molecular basis behind the stereoselectivity difference, av ariety of aromatick etones and b-ketoesters were docked into the active center of BaSDR1 (Figure 1a nd Figures S1-S10 in the Supporting Information).I n our previous studies, the mechanisms regarding the catalytic performance of BaSDR1 against aromatic ketones have been deeply discussed. [15,22] Herein, the complex of BaSDR1 and sub-Scheme1.Asymmetric reduction of prochiral ketones to the corresponding chiral alcohols by BaSDR1. strate 23 a was chosen as ar epresentativem odel of aromatic ketones to avoid unnecessaryr epetition.…”

Section: Resultsmentioning

confidence: 99%

“…The three‐dimensional homology model of wildtype BaSDR1 was constructed by using Modeller as in our previous report [15, 22] . The structure models of mutants were generated by using Swiss‐PDB Viewer.…”

Section: Methodsmentioning

confidence: 99%

“…Recombinant Escherichia coli BL21(DE3) strain harboring pET30a-BaSDR1 has been constructed in our previous study. [15] Variants were generated by PCR-based site-directed mutagenesis using the recombinant plasmid pET30a-BaSDR1 as the template and the corresponding primers are listed in Ta ble S1 (in the Supporting Information). The PCR was carried out by established methods.…”

Section: Mutant Constructionmentioning

confidence: 99%

“…In previous work, a carbonyl reductase (BaSDR1) from Bacillus aryabhattai was discovered by the genome hunting strategy [15] . It was highly active for the reduction of aromatic ketones, especially ortho ‐haloacetophenones, giving the corresponding ( S )‐alcohols with 99.9 % enantiomeric excess ( ee ).…”

Section: Introductionmentioning

confidence: 99%

“…In previous work, ac arbonyl reductase (BaSDR1) from Bacillus aryabhattai was discovered by the genomeh unting strategy. [15] It was highly active for the reduction of aromatic ketones, especially ortho-haloacetophenones, giving the corresponding (S)-alcohols with 99.9 %e nantiomeric excess( ee). However,B aSDR1e xhibitedamoderate stereoselectivity toward b-ketoesters.T he ee value of corresponding b-hydroxy esters varied from 55.9 %t o9 7.0 %( Scheme 1).…”

Section: Introductionmentioning

confidence: 99%

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Single‐Point Mutant Inverts the Stereoselectivity of a Carbonyl Reductase toward β‐Ketoesters with Enhanced Activity

Wang

Tian

et al. 2021

Chemistry A European J

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Enzyme stereoselectivity control is still a major challenge. To gain insight into the molecular basis of enzyme stereo‐recognition and expand the source of antiPrelog carbonyl reductase toward β‐ketoesters, rational enzyme design aiming at stereoselectivity inversion was performed. The designed variant Q139G switched the enzyme stereoselectivity toward β‐ketoesters from Prelog to antiPrelog, providing corresponding alcohols in high enantiomeric purity (89.1–99.1 % ee). More importantly, the well‐known trade‐off between stereoselectivity and activity was not found. Q139G exhibited higher catalytic activity than the wildtype enzyme, the enhancement of the catalytic efficiency (kcat/Km) varied from 1.1‐ to 27.1‐fold. Interestingly, the mutant Q139G did not lead to reversed stereoselectivity toward aromatic ketones. Analysis of enzyme–substrate complexes showed that the structural flexibility of β‐ketoesters and a newly formed cave together facilitated the formation of the antiPrelog‐preferred conformation. In contrast, the relatively large and rigid structure of the aromatic ketones prevents them from forming the antiPrelog‐preferred conformation.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Mutant Constructionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

Single‐Point Mutant Inverts the Stereoselectivity of a Carbonyl Reductase toward β‐Ketoesters with Enhanced Activity

Wang

Tian

et al. 2021

Chemistry A European J

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Engineering a Carbonyl Reductase as a Potential Tool for the Synthesis of Chiral α‐Tetralinols

Wang

Yang

et al. 2021

ChemCatChem

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Tailoring of enzyme toward α‐tetralones, a class of bulky‐bulky ketones, is still a challenge. In this work, the mutants of carbonyl reductase BaSDR1 with improved catalytic performance toward α‐tetralone 1 a were obtained by adjusting the steric hindrance and hydrophobicity of the residues that affect the approach of α‐tetralone with the catalytic residues. The designed mutants also showed enhanced catalytic performance toward halogenated α‐tetralones 2 a–6 a. Remarkably, the activity of the mutant Q237V/I291F toward 7‐fluoro‐α‐tetralone 5 a was 16.3‐fold higher than the wildtype enzyme with improved stereoselectivity (98.8 % ee). More notably, the mutants Q139S and Q139S/V187S exhibited decreased or reversed stereoselectivity toward α‐tetralone 1 a, 5‐bromo‐α‐tetralone 2 a, 7‐fluoro‐α‐tetralone 5 a and 7‐chloro‐α‐tetralone 6 a, while the relatively high ee values were obtained in the presence of 6‐chloro‐α‐tetralone 3 a and 6‐bromo‐α‐tetralone 4 a as substrates. Further analysis showed the larger size of the substrates was beneficial for the substrates binding to the active cavity with a more specific binding mode, which endows the reaction with higher stereoselectivity. Moreover, the recombinant E. coli expressing the variant Q237V/I291F successfully catalyzed the reduction of a high concentration 7‐fluoro‐α‐tetralone 5 a. These results not only offered a potential tool for chiral α‐tetralols, but also provided guiding information for the enzyme engineering toward bulky‐bulky ketones.

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Recent advances in biocatalytic derivatization of l-tyrosine

Tan

Song

Chen

et al. 2020

Appl Microbiol Biotechnol

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Discovery of a novel ortho-haloacetophenones-specific carbonyl reductase from Bacillus aryabhattai and insight into the molecular basis for its catalytic performance

Cited by 13 publications

References 57 publications

Single‐Point Mutant Inverts the Stereoselectivity of a Carbonyl Reductase toward β‐Ketoesters with Enhanced Activity

Single‐Point Mutant Inverts the Stereoselectivity of a Carbonyl Reductase toward β‐Ketoesters with Enhanced Activity

Engineering a Carbonyl Reductase as a Potential Tool for the Synthesis of Chiral α‐Tetralinols

Recent advances in biocatalytic derivatization of l-tyrosine

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