College of Chemical Engineering, Huaqiao University, 668 Jimei Blvd., Xiamen, Fujian 361021, China; School of Environmental Science and Engineering, Shandong University, Qingdao 266237, China.
College of Chemical Engineering, Huaqiao University, 668 Jimei Blvd., Xiamen, Fujian 361021, China.
Enzyme Microb Technol. 2022 Oct;160:110074. doi: 10.1016/j.enzmictec.2022.110074. Epub 2022 Jun 7.
In recent years, with the increasing number of patients with depression, the efficient synthesis of the first-line antidepressant drug duloxetine intermediate (S-N,N-dimethyl-3-hydroxy-3-(2-thienyl)-1-propanamine, S-DHTP) has attracted great attention. The wild-type AKR3-2-9 from Bacillus megaterium YC4-R4 exhibits high application potential of catalyzing N,N-dimethyl-3-keto-3-(2-thienyl)-1-propanamine (DKTP) to prepare S-DHTP, but there is still much room for improvement. In this work, rational design was carried out to enhance the catalytic potential of AKR3-2-9. Notably, compared to the wild-type AKR3-2-9, three mutants (Ile189Val, Asn256Asp, and Ile189Val + Asn256Asp) were obtained, and their catalytic efficiencies were increased by 1.3 times, 2.3 times, and 1.31 times, respectively. Besides, the thermal stability and organic solvent resistance were improved. More importantly, when the concentration of the substrate DKTP was 0.5 g/L, the catalytic yields of Ile189Val, Asn256Asp and Ile189Val + Asn256Asp were increased by 1.45 times, 1.86 times, and 2.05 times, respectively. Besides, the corresponding optical purities of the three mutants were 92.7 %, 94.3 % and 93.8 %. The above results indicated that the rational design of the AKR of Bacillus megaterium YC4-R4 enhanced its potential for biocatalytic preparation of S-DHTP.
近年来，随着抑郁症患者数量的增加，高效合成一线抗抑郁药度洛西汀中间体（S-N,N-二甲基-3-羟基-3-（2-噻吩基）-1-丙胺，S-DHTP）引起了极大的关注。来自巨大芽孢杆菌 YC4-R4 的野生型 AKR3-2-9 显示出催化 N,N-二甲基-3-酮-3-（2-噻吩基）-1-丙胺（DKTP）制备 S-DHTP 的高应用潜力，但仍有很大的改进空间。在这项工作中，进行了合理的设计以增强 AKR3-2-9 的催化潜力。值得注意的是，与野生型 AKR3-2-9 相比，获得了三个突变体（Ile189Val、Asn256Asp 和 Ile189Val+Asn256Asp），它们的催化效率分别提高了 1.3 倍、2.3 倍和 1.31 倍。此外，还提高了热稳定性和有机溶剂耐受性。更重要的是，当底物 DKTP 的浓度为 0.5 g/L 时，Ile189Val、Asn256Asp 和 Ile189Val+Asn256Asp 的催化产率分别提高了 1.45 倍、1.86 倍和 2.05 倍。此外，这三种突变体的相应光学纯度分别为 92.7%、94.3%和 93.8%。上述结果表明，对巨大芽孢杆菌 YC4-R4 的 AKR 进行合理设计增强了其生物催化制备 S-DHTP 的潜力。
