Journal of Microbiology and Biotechnology
The Korean Society for Microbiology and Biotechnology publishes the Journal of Microbiology and Biotechnology.

2020 ; Vol.30-1: 146~154

AuthorWei Jiang, Yali Wang
Place of dutyDepartment of Bioengineering and Biotechnology, College of Chemical Engineering, Huaqiao University, Xiamen, Fujian, 361021, P.R. China
TitleImproving Catalytic Efficiency and Changing Substrate Spectrum for Asymmetric Biocatalytic Reductive Amination
PublicationInfo J. Microbiol. Biotechnol.2020 ; Vol.30-1
AbstractWith the advantages of biocatalytic method, enzymes have been excavated for the synthesis of chiral amino acids by the reductive amination of ketones, offering a promising way of producing pharmaceutical intermediates. In this work, a robust phenylalanine dehydrogenase (PheDH) with wide substrate spectrum and high catalytic efficiency was constructed through rational design and active-site-targeted, site-specific mutagenesis by using the parent enzyme from Bacillus halodurans. Active sites with bonding substrate and amino acid residues surrounding the substrate binding pocket, 49L-50G-51G, 74M,77K, 122G-123T-124D-125M, 275N, 305L and 308V of the PheDH, were identified. Noticeably, the new mutant PheDH (E113D-N276L) showed approximately 6.06-fold increment of kcat/Km in the oxidative deamination and more than 1.58-fold in the reductive amination compared to that of the wide type. Meanwhile, the PheDHs exhibit high capacity of accepting benzylic and aliphatic ketone substrates. The broad specificity, high catalytic efficiency and selectivity, along with excellent thermal stability, render these broad-spectrum enzymes ideal targets for further development with potential diagnostic reagent and pharmaceutical compounds applications.
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Key_wordChiral compounds, asymmetric catalysis, substrate spectrum, biocatalysis, rational design, substrate binding pocket
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