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Improving Catalytic Efficiency and Changing Substrate Spectrum for Asymmetric Biocatalytic Reductive Amination
1Department of Bioengineering and Biotechnology, College of Chemical Engineering, Huaqiao University, Xiamen, Fujian, 361021, P.R. China.
Journal of Microbiology and Biotechnology
|September 24, 2019
Summary
Engineered phenylalanine dehydrogenase (PheDH) enzymes offer a greener route for synthesizing chiral amino acids from ketones. Mutants show enhanced efficiency and broad substrate acceptance for pharmaceutical applications.
Area of Science:
- Biocatalysis and Enzyme Engineering
- Organic Synthesis
- Pharmaceutical Chemistry
Background:
- Enzymes are valuable biocatalysts for synthesizing chiral amino acids, crucial pharmaceutical intermediates.
- Reductive amination of ketones using enzymes provides a sustainable synthetic approach.
- Phenylalanine dehydrogenase (PheDH) is a key enzyme in amino acid metabolism.
Purpose of the Study:
- To engineer a robust phenylalanine dehydrogenase (PheDH) with an expanded substrate spectrum and improved catalytic efficiency.
- To identify key amino acid residues in the active site of PheDH that influence substrate binding and catalysis.
- To develop novel biocatalysts for the efficient production of chiral amino acids.
Main Methods:
- Rational enzyme design and site-specific mutagenesis were employed to modify the parent PheDH from *Bacillus halodurans*.
- Active site residues involved in substrate binding were identified through structural analysis.
- Enzyme activity was assessed for both oxidative deamination and reductive amination reactions.
Main Results:
- A mutant PheDH (E113D-N276L) exhibited a 6.06-fold increase in catalytic rate for oxidative deamination and a 1.58-fold increase for reductive amination.
- The engineered PheDH demonstrated broad substrate specificity, accepting both benzylic and aliphatic ketones.
- The enzyme displayed high catalytic efficiency, selectivity, and enhanced thermal stability.
Conclusions:
- Engineered PheDH enzymes represent a significant advancement in biocatalytic synthesis of chiral amino acids.
- The developed broad-spectrum enzymes hold potential for applications as diagnostic reagents and in pharmaceutical compound development.
- Rational design and mutagenesis are effective strategies for enhancing enzyme performance and expanding their utility.
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