Creation of a Broad-Range and Highly Stereoselective D-Amino Acid Dehydrogenase for the One-Step Synthesis of D-Amino Acids

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• 作者：Kavitha Vedha-Peters ; Manjula Gunawardana ; J. David Rozzell ; Scott J. Novick
• 刊名：Journal of the American Chemical Society
• 出版年：2006
• 出版时间：August 23, 2006
• 年：2006
• 卷：128
• 期：33
• 页码：10923 - 10929
• 全文大小：98K
• 年卷期：v.128,no.33(August 23, 2006)
• ISSN：1520-5126

文摘

Using both rational and random mutagenesis, we have created the first known broad substraterange, nicotinamide cofactor dependent, and highly stereoselective D-amino acid dehydrogenase. Thisnew enzyme is capable of producing D-amino acids via the reductive amination of the corresponding 2-ketoacid with ammonia. This biocatalyst was the result of three rounds of mutagenesis and screening performedon the enzyme meso-diaminopimelate D-dehydrogenase. The first round targeted the active site of thewild-type enzyme and produced mutants that were no longer strictly dependent on the native substrate.The second and third rounds produced mutants that had an increased substrate range including straight-and branched-aliphatic amino acids and aromatic amino acids. The very high selectivity toward theD-enantiomer (95 to >99% ee) was shown to be preserved even after the addition of the five mutationsfound in the three rounds of mutagenesis and screening. This new enzyme could complement and improveupon current methods for D-amino acid synthesis.