Fengyu Kou1, Jing Zhao1, Jiao Liu1, Cunmin Sun1, Yanmei Guo1, Zijian Tan1, Feng Cheng2, Zhimin Li3, Ping Zheng4, Jibin Sun1. 1. Key Laboratory of Systems Microbial Biotechnology, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin, 300308, China. 2. College of Biotechnology and Bioengineering, Zhejiang University of Technology, Hangzhou, 310014, China. 3. State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, 200237, China. 4. Key Laboratory of Systems Microbial Biotechnology, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin, 300308, China. zheng_p@tib.cas.cn.
Abstract
OBJECTIVE: To enhance the thermal and alkaline pH stability of the lysine decarboxylase from Escherichia coli (CadA) by engineering the decameric interface and explore its potential for industrial applications. RESULTS: The mutant T88S was designed for improved structural stability by computational analysis. The optimal pH and temperature of T88S were 7.0 and 55 °C (5.5 and 50 °C for wild-type). T88S showed higher thermostability with a 2.9-fold increase in the half-life at 70 °C (from 11 to 32 min) and increased melting temperature (from 76 to 78 °C). Additionally, the specific activity and pH stability (residual activity after 10 h incubation) of T88S at pH 8.0 were increased to 164 U/mg and 78% (58 U/mg and 57% for wild-type). The productivity of cadaverine with T88S (284 g L-lysine L-1 and 5 g DCW L-1) was 40 g L-1 h-1, in contrast to 28 g L-1 h-1 with wild-type. CONCLUSION: The mutant T88S showed high thermostability, pH stability, and activity at alkaline pH, indicating that this mutant is a promising biocatalyst for industrial production of cadaverine.
OBJECTIVE: To enhance the thermal and alkaline pH stability of the lysine decarboxylase from Escherichia coli (CadA) by engineering the decameric interface and explore its potential for industrial applications. RESULTS: The mutant T88S was designed for improved structural stability by computational analysis. The optimal pH and temperature of T88S were 7.0 and 55 °C (5.5 and 50 °C for wild-type). T88S showed higher thermostability with a 2.9-fold increase in the half-life at 70 °C (from 11 to 32 min) and increased melting temperature (from 76 to 78 °C). Additionally, the specific activity and pH stability (residual activity after 10 h incubation) of T88S at pH 8.0 were increased to 164 U/mg and 78% (58 U/mg and 57% for wild-type). The productivity of cadaverine with T88S (284 g L-lysine L-1 and 5 g DCW L-1) was 40 g L-1 h-1, in contrast to 28 g L-1 h-1 with wild-type. CONCLUSION: The mutant T88S showed high thermostability, pH stability, and activity at alkaline pH, indicating that this mutant is a promising biocatalyst for industrial production of cadaverine.