Fine-tuning of the substrate binding mode to enhance the catalytic efficiency of an ortho-haloacetophenone-specific carbonyl reductase. Issue 8 (23rd March 2020)

• Record Type: Journal Article
• Authors: Li, Aipeng; Li, Xue; Pang, Wei; Tian, Qing; Wang, Ting; Zhang, Lianbing
• Is Part Of: Catalysis science & technology. Volume 10:Issue 8(2020)
• Page Start: 2462
• Page End: 2472
• Publication Date: 2020-03-23
• Subjects: Catalysis -- Periodicals; 541.395
• DOI: 10.1039/c9cy02335f
• Languages: English
• ISSNs: 2044-4753

Abstract

Abstract : Fine-tuning of the substrate binding mode was successfully applied for enhancing the catalytic efficiency of an ortho -haloacetophenone-specific carbonyl reductase. Abstract : Carbonyl reductase BaSDR1 has been identified as a potential ortho -haloacetophenone-specific biocatalyst for the synthesis of chiral 1-(2-halophenyl)ethanols due to its excellent stereoselectivity. However, the catalytic efficiency of BaSDR1 is far below the required level for practical applications. Thus, fine-tuning of the substrate binding mode, which aimed at maximum preservation of the positive factors for substrate specificity and stereoselectivity, was proposed as a tentative strategy for enhancing its catalytic efficiency. The designed mutants Q139S, D253Y and Q139S/D253Y showed significantly enhanced catalytic efficiency. Remarkably, the variants Q139S and Q139S/D253Y exhibited a more than 9-fold improvement in catalytic efficiency ( k cat / K m ) toward substrates 6a and 11a, respectively. More importantly, none of the variants caused activity–stereoselectivity trade-off and all variants exhibited excellent stereoselectivity (99% ee). Analysis of variant–substrate complexes showed that the mutations indeed enable the fine-tuning of the substrate binding mode. New strengthening factors for consolidating the productive conformation were introduced while the original positive factors were preserved. Furthermore, at a substrate concentration of 100 mM, recombinant E. coli whole cells expressing the BaSDR1 mutants were successfully applied to the synthesis of several key intermediates of chiral pharmaceuticals, including ( S )-1-(2-chlorophenyl)ethanol, ( S )-1-(2, 4-difluorophenyl)ethanol and ( S )-1-(2, 6-difluorophenyl)ethanol, with 99% enantiomeric excess, and the conversion reached over 95% in a certain period of time. These results demonstrated the effectiveness of the strategy involving the fine-tuning of the substrate binding mode and the applicability of the designed mutants in efficient reduction of ortho -haloacetophenones.