1. Directed Evolution Improves the Enzymatic Synthesis of L-5-Hydroxytryptophan by an Engineered Tryptophan Synthase.
- Author
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Xu L, Li T, Huo Z, Chen Q, Xia Q, and Jiang B
- Subjects
- Protein Engineering methods, Tryptophan Synthase genetics, Tryptophan Synthase metabolism, Tryptophan Synthase chemistry, Directed Molecular Evolution, Escherichia coli genetics, 5-Hydroxytryptophan biosynthesis
- Abstract
L-5-Hydroxytryptophan is an important amino acid that is widely used in food and medicine. In this study, L-5-hydroxytryptophan was synthesized by a modified tryptophan synthase. A direct evolution strategy was applied to engineer tryptophan synthase from Escherichia coli to improve the efficiency of L-5-hydroxytryptophan synthesis. Tryptophan synthase was modified by error-prone PCR. A high-activity mutant enzyme (V231A/K382G) was obtained by a high-throughput screening method. The activity of mutant enzyme (V231A/K382G) is 3.79 times higher than that of its parent, and k
cat /Km of the mutant enzyme (V231A/K382G) is 4.36 mM-1 ∙s-1 . The mutant enzyme (V231A/K382G) reaction conditions for the production of L-5-hydroxytryptophan were 100 mmol/L L-serine at pH 8.5 and 35°C for 15 h, reaching a yield of L-5-hydroxytryptophan of 86.7%. Directed evolution is an effective strategy to increase the activity of tryptophan synthase., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)- Published
- 2021
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