Resolution Mechanism and Characterization of an Ammonium Chloride-Tolerant, High-Thermostable, and Salt-Tolerant Phenylalanine Dehydrogenase from Bacillus halodurans
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As phenylalanine dehydrogenase (PheDH) plays an important role in the synthesis of chiral drug intermediates and detection of phenylketonuria, it is significant to obtain a PheDH with specific and high activity. Here, a PheDH gene, pdh, encoding a novel BhPheDH with 61.0% similarity to the known PheDH from Microbacterium sp., was obtained. The BhPheDH showed optimal activity at 60 °C and pH 7.0, and it showed better stability in hot environment (40–70 °C) than the PheDH from Nocardia sp. And its activity and thermostability could be significantly increased by sodium salt. After incubation for 2 h in 3 M NaCl at 60 °C, the residual activity of the BhPheDH was found to be 1.8-fold higher than that of the control group (without NaCl). The BhPheDH could tolerate high concentration of ammonium chloride and its activity could be also enhanced by the high concentration of ammonium salts. These characteristics indicate that the BhPheDH possesses better thermostability, ammonium chloride tolerance, halophilic mechanism, and high salt activation. The mechanism of thermostability and high salt tolerance of the BhPheDH was analyzed by molecular dynamics simulation. These results provide useful information about the enzyme with high-temperature activity, thermostability, halophilic mechanism, tolerance to high concentration of ammonium chloride, higher salt activation and enantio-selectivity, and the application of molecular dynamics simulation in analyzing the mechanism of these distinctive characteristics.
KeywordsThermostability Mechanism of salt tolerance Phenylalanine dehydrogenase Molecular dynamics simulation Chiral drug intermediates
WJ designed and performed the experiments and wrote the manuscript. YZW performed the experiments and revised the manuscript. BSF conceived the study, designed and supervised the experiments, and is a corresponding author. All authors have read and approved the manuscript.
This work was supported by the State Key Program of National Natural Science Foundation of China (No. 21336009), the National Natural Science Foundation of China (No. 41176111, No. 41306124), the Foundation of South Oceanographic Research Center of China in Xiamen (No.: 14GYY011NF11), the Public science and technology research funds projects of ocean (No.: 201505032-6), and the high-level personnel activation fee of Huaqiao University (600005-Z17Y0072).
Compliance with Ethical Standards
Conflict of Interest
The authors declare that they have no competing interests.
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