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Redesign of a Phenylalanine Aminomutase into a Phenylalanine Ammonia Lyase

Authors

  • Dr. Sebastian Bartsch,

    1. Department of Biochemistry, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Nijenborgh 4, 9747 AG, Groningen (The Netherlands)
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  • Gjalt G. Wybenga,

    1. Laboratory of Biophysical Chemistry, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen (The Netherlands)
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  • Maaike Jansen,

    1. Department of Biochemistry, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Nijenborgh 4, 9747 AG, Groningen (The Netherlands)
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  • Matthew M. Heberling,

    1. Department of Biochemistry, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Nijenborgh 4, 9747 AG, Groningen (The Netherlands)
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  • Dr. Bian Wu,

    1. Department of Biochemistry, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Nijenborgh 4, 9747 AG, Groningen (The Netherlands)
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  • Prof. Bauke W. Dijkstra,

    1. Laboratory of Biophysical Chemistry, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen (The Netherlands)
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  • Prof. Dick B. Janssen

    Corresponding author
    1. Department of Biochemistry, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Nijenborgh 4, 9747 AG, Groningen (The Netherlands)
    • Department of Biochemistry, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Nijenborgh 4, 9747 AG, Groningen (The Netherlands)

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Abstract

An aminomutase, naturally catalyzing the interconversion of (S)-α-phenylalanine and (R)-β-phenylalanine, was converted into an ammonia lyase catalyzing the nonoxidative deamination of phenylalanine to cinnamic acid by a rational single-point mutation. It could be shown by crystal structures and kinetic data that the flexibility of the lid that covers the active site decides whether the enzyme acts as a lyase or a mutase. An Arg92Ser mutation destabilized the closed conformation of the lid structure and converted the mutase into a lyase that exhibited up to 44-fold increased reaction rates in the enantioselective deamination of (R)-β-phenylalanine. In addition, the amination rates of cinnamic acid yielding optically pure (S)-α- and (R)-β-phenylalanine were doubled. The applicability of the mutant enzyme for kinetic resolution and asymmetric amination could be shown by biocatalysis on a preparative scale.

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