Abstract
Glycerol dehydratase (GDHt) is a key and rate-limiting enzyme in the pathway of 1,3-propanediol (1,3-PD) synthesis. The improvement of GDHt’s stability and enzymatic activity is desirable for the biosynthesis of 1,3-PD. The gldABC gene encoding GDHt of Klebsiella pneumoniae was cloned and expressed in Escherichia coli XL10-Gold, and the mutation sites of GDHt were obtained through prediction by PoPMuSiC program. Consequently, two mutants (KpG60 and KpG525) were developed by rational design through site-mutagenesis based on 3D structure which was constructed from homology modeling. Analyses of enzymatic properties showed that pH stability of the mutants was about 1.25–2 times higher than that of the wild type, and specific activity, Vmax and Kcat/Km of KpG525 were about 1.5–2 times higher than those of the wild type. This work presented a simple and useful measure to improve the performance of industrial enzyme.
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Acknowledgments
We thank Prof. Yong Wang of Jiangsu University for his revision to the manuscript. This work was supported by the National Natural Science Foundation of China under Grant No. 21006041, the Postdoctoral Foundation of Jiangsu Province under Grant No. 0901012B and the Scientific Research Promotion Fund for the Talents of Jiangsu University under Grant No. 08JDG009.
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Qi, X., Guo, Q., Wei, Y. et al. Enhancement of pH stability and activity of glycerol dehydratase from Klebsiella pneumoniae by rational design. Biotechnol Lett 34, 339–346 (2012). https://doi.org/10.1007/s10529-011-0775-5
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DOI: https://doi.org/10.1007/s10529-011-0775-5