Modeling and Optimization of Phenylalanine Ammonia Lyase Stabilization in Recombinant Escherichia coli for the Continuous Synthesis of l-Phenylalanine on the Statistical-Based Experimental Designs

Some approaches for improving recombinant phenylalanine ammonia lyase (PAL) stability in Escherichia coli during the enzymatic methods of l-phenylalanine (l-Phe) production were developed following preliminary studies by means of statistical-based experiment designs (response surface method). The tr...

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Bibliographic Details
Published in:Journal of agricultural and food chemistry 2010-03, Vol.58 (5), p.2795-2800
Main Authors: Zhang, Bing-Zhu, Cui, Jian-Dong, Zhao, Gui-Xia, Jia, Shi-Ru
Format: Article
Language:English
Subjects:
Analytical Methods
biotechnology
biotransformation
continuous systems
enzyme activity
Enzyme Stability
Escherichia coli
Escherichia coli - genetics
Models, Theoretical
phenylalanine
Phenylalanine - metabolism
phenylalanine ammonia-lyase
Phenylalanine Ammonia-Lyase - metabolism
recombinant DNA
Recombinant Proteins - metabolism
statistical models
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Summary:Some approaches for improving recombinant phenylalanine ammonia lyase (PAL) stability in Escherichia coli during the enzymatic methods of l-phenylalanine (l-Phe) production were developed following preliminary studies by means of statistical-based experiment designs (response surface method). The traditional non-statistical technology was used to screen four critical factors for PAL stability during the bioconversion process, viz., glycerin, sucrose, 1,4-dithiothreitol (DTT), and MgSO4. The central composite design (CCD) was applied to optimize the combined effect of critical factors for recombinant PAL stability and understand the relationship between the factors and PAL stability. The optimum values for testing variables were 13.04 mM glycerin, 1.87 mM sucrose, 4.09 mM DTT, and 69 mM Mg2+. A second-order model equation was suggested and then validated experimentally. The model adequacy was very satisfactory because the coefficient of determination was 0.88. The maximum PAL activity was retained as 67.73 units/g after three successive cycles of bioconversion. In comparison to initial PAL activity, the loss of PAL activity was only 22%. PAL activity was enhanced about 23% in comparison to the control (without any stabilizer additives). PAL stability was significantly improved during successive bioconversion. The results obtained here verified the effectiveness of the applied methodology and may be helpful for l-Phe production on an industrial scale.
ISSN:0021-8561
1520-5118
DOI:10.1021/jf9036744