National and Local Joint Engineering Research Center for Biomanufacturing of Chiral Chemicals, Zhejiang University of Technology, Hangzhou 310014, People's Republic of China; Key Laboratory of Bioorganic Synthesis of Zhejiang Province, College of Biotechnology and Bioengineering, Zhejiang University of Technology, Hangzhou 310014, People's Republic of China.
National and Local Joint Engineering Research Center for Biomanufacturing of Chiral Chemicals, Zhejiang University of Technology, Hangzhou 310014, People's Republic of China; Key Laboratory of Bioorganic Synthesis of Zhejiang Province, College of Biotechnology and Bioengineering, Zhejiang University of Technology, Hangzhou 310014, People's Republic of China.
Bioorg Chem. 2023 Nov;140:106788. doi: 10.1016/j.bioorg.2023.106788. Epub 2023 Aug 14.
Vibegron is a novel, potent, highly selective β-adrenergic receptor agonist for the treatment of overactive bladder with higher therapeutic capacity and lower side effects. Methyl(2S,3R)-2-((tert-butoxycarbonyl)amino)-3-hydroxy-3-phenylpropanoate ((2S,3R)-aminohydroxy ester) is a key chiral intermediate for the synthesis of Vibegron. A novel carbonyl reductase from Exiguobacterium sp. s126 (EaSDR6) was isolated using data mining technology from GenBank database with preferable catalytic activity. Hydrogen bond network regulation was performed using site-directed saturation mutagenesis and combination mutagenesis. The mutant EaSDR6 was obtained with the activity improvement by 4.58 folds compared with the wild type EaSDR6. The K of EaSDR6 was decreased from 1.57 mM to 0.67 mM, k was increased by 2.17 folds, and the overall catalytic efficiency k/K was increased by 5.07 folds. The organic-aqueous biphasic bioreaction system for the asymmetric synthesis of (2S,3R)-aminohydroxy ester was constructed for the first time. Under the substrate concentration of 150 g/L, the yield of (2S,3R)-aminohydroxy ester was > 99.99%, the e.e. was > 99.99%, and the spatiotemporal yield was 1.55 g/(L·h·g ) after 12 h reaction. While the substrate concentration was increased to 200 g/L and the reaction lasted for 36 h, the yield of (2S,3R)-aminohydroxy ester was > 99.99%, the e.e. was > 99.99% and the spatiotemporal yield was 1.05 g/(L·h·g ). The substrate concentration and spatiotemporal yield were higher than ever reported.
Vibegron 是一种新型、高效、高选择性的β-肾上腺素能受体激动剂，用于治疗膀胱过度活动症，具有更高的治疗效果和更低的副作用。(2S,3R)-2-((叔丁氧基羰基)氨基)-3-羟基-3-苯基丙酸甲酯((2S,3R)-氨基羟酯)是 Vibegron 合成的关键手性中间体。利用 GenBank 数据库中的数据挖掘技术，从极端微生物 Exiguobacterium sp. s126 中分离到一种新型的羰基还原酶(EaSDR6)，其具有较好的催化活性。通过定点饱和突变和组合突变进行氢键网络调控。与野生型 EaSDR6 相比，突变体 EaSDR6 的活性提高了 4.58 倍。EaSDR6 的 K 值从 1.57mM 降低到 0.67mM，k 值提高了 2.17 倍，总催化效率 k/K 提高了 5.07 倍。首次构建了用于(2S,3R)-氨基羟酯不对称合成的有机-水两相生物反应体系。在底物浓度为 150g/L 的条件下，(2S,3R)-氨基羟酯的产率>99.99%，ee 值>99.99%，时空产率为 1.55g/(L·h·g)，反应 12h 后。当底物浓度增加到 200g/L，反应持续 36h 时，(2S,3R)-氨基羟酯的产率>99.99%，ee 值>99.99%，时空产率为 1.05g/(L·h·g)。底物浓度和时空产率均高于以往报道。
