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KMID : 0545120190290050749
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Journal of Microbiology and Biotechnology
2019 Volume.29 No. 5 p.749 ~ p.757
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Production of Cyanocarboxylic Acid by Acidovorax facilis 72W Nitrilase Displayed on the Spore Surface of Bacillus subtilis
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Zhong Xia
Yang Shaomin
Su Xinying
Shen Xiaoxia
Zhao Wen
Chan Zhi
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Abstract
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Nitrilase is a valuable type of hydrolase that catalyzes nitriles into carboxylic acid and ammonia. Its applications, however, are severely restricted by the harsh conditions of industrial reaction processes. To solve this problem, a nitrilase from Acidovorax facilis 72W was inserted into an Escherichia coli?Bacillus subtilis shuttle vector for spore surface display. Western blot, enzyme activity measurements and flow cytometric analysis results all indicated a successful spore surface display of the CotB-nit fusion protein. In addition, the optimal catalytic pH value and temperature of the displayed nitrilase were determined to be 7.0 and 50°C, respectively. Moreover, results of reusability tests revealed that 64% of the initial activity of the displayed nitrilase was still retained at the 10th cycle. Furthermore, hydrolysis efficiency of upscale production of cyanocarboxylic acid was significantly higher in the displayed nitrilase-treated group than in the free group expressed by E. coli (pET-28a-nit). Generally, the display of A. facilis 72W nitrilase on the spore surface of Bacillus subtilis may be a useful method for immobilization of enzyme and consequent biocatalytic stabilization.
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KEYWORD
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Acidovorax facilis 72W, nitrilase, Bacillus subtilis, immobilization, spore surface display
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