Combining Binding Pocket Mutagenesis and Substrate Tunnel Engineering to Improve an (R)-Selective Transaminase for the Efficient Synthesis of (R)-3-aminobutanol
Biochemical and biophysical research communications(2024)
摘要
(R)-selective transaminases have the potential to act as efficient biocatalysts for the synthesis of important pharmaceutical intermediates. However, their low catalytic efficiency and unfavorable equilibrium limit their industrial application. Seven (R)-selective transaminases were identified using homologous sequence mining. Beginning with the optimal candidate from Mycolicibacterium hippocampi, virtual mutagenesis and substrate tunnel engineering were performed to improve catalytic efficiency. The obtained variant, T282S/Q137E, exhibited 3.68-fold greater catalytic efficiency (kcat/Km) than the wild-type enzyme. Using substrate fed-batch and air sweeping processes, effective conversion of 100 mM 4-hydroxy-2-butanone was achieved with a conversion rate of 93 % and an ee value > 99.9 %. This study provides a basis for mutation of (R)-selective transaminases and offers an efficient biocatalytic process for the asymmetric synthesis of (R)-3-aminobutanol.
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关键词
(R ) -Selective transaminase,Asymmetric synthesis,Virtual mutagenesis,Tunnel engineering,Fed -batch
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