Stereoselective synthesis of l-tert-leucine by a newly cloned leucine dehydrogenase from Exiguobacterium sibiricum
2014
Li, Jing | Pan, Jiang | Zhang, Jie | Xu, Jian-He
A leucine dehydrogenase from Exiguobacterium sibiricum (EsLeuDH) was discovered by genome mining approach. The EsLeuDH was overexpressed in Escherichia coli BL21, purified to homogeneity and characterized. This enzyme showed good thermostability with a half-life of 3.1h at 60°C. Furthermore, EsLeuDH has a broad spectrum of substrate specificity, showing activities toward many aliphatic α-keto acids and L-amino acids, in addition to some aryl α-keto acids and aryl α-amino acids, such as α-oxobenzeneacetic and l-phenylglycine. The EsLeuDH was successfully coexpressed with Bacillus megaterium glucose dehydrogenase (BmGDH) in Escherichia coli BL21 for the production of l-tert-leucine. By using the coexpressed whole cells, a decagram preparation of l-tert-leucine was performed at a substrate concentration of 0.6M (78.1gL−1) in 1L scale with 99% conversion after 5.5h, resulting in 80.1% yield and>99% ee (enantiomeric excess).
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