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Immobilization of phenylalanine dehydrogenase onto Eupergit CM for the synthesis of (S)-2-amino-4-phenylbutyric acid

Ahmad, A.L., Low, E.M., Shukor, S.R. Abd
Journal of Molecular Catalysis. B, Enzymatic 2013 v.88 pp. 26-31
catalytic activity, pH, phenylalanine, storage quality, temperature, thermal stability
An oxidoreductase enzyme, phenylalanine dehydrogenase (PheDH) from Rhodoccocus sp. 4 was covalently immobilized onto a commercially available enzyme carrier, Eupergit CM and tested for the synthesis of an unnatural amino acid, (S)-2-amino-4-phenylbutyric acid (S-APBA). Through performance evaluation of the immobilization based on its yield and efficiency, an adequate combination of the immobilization conditions to obtain optimum values in these parameters were determined as enzyme loading of 60:1 (weight ratio of enzyme carrier to enzyme) in an immobilization buffer of 500mM at pH 8.5 for 24h. Upon covalent immobilization onto Eupergit CM, the operational pH range for reductive amination was slightly broadened (pH 7.5–9.5). Immobilization also helps to improve the thermostability of PheDH, raising the optimum temperature to 50°C. A remarkable improvement in operability and storage stability was achieved. The immobilized PheDH was successfully applied for the synthesis of (S)-2-amino-4-phenylbutyric acid, achieving enantiomeric excess of more than 99% and yield of more than 80%; comparable to synthesis using the free PheDH. Reductive amination using the immobilized PheDH is beneficial to enhance the asymmetric synthesis S-APBA.