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KMID : 1094720130180020358
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Biotechnology and Bioprocess Engineering
2013 Volume.18 No. 2 p.358 ~ p.364
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Enzymatic synthesis of theanine with Escherichia coli γ-glutamyltranspeptidase from a series of γ-glutamyl anilide substrate analogues
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Zhang Hong-Juan
Zhang Wei-Guo
Wang Zhi-Yuan
Zhan Yue-Ping
Xu Li-Sheng
Liu Jun-Zhong
Liu Qian
Jiao Qing-Cai
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Abstract
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In order to investigate the catalytic mechanism of Escherichia coli γ-glutamyltranspeptidase, ten para- and meta-substituted γ-glutamyl anilides were chemically prepared and employed as substrates to synthesize L-theanine to assay the activity of γ-glutamyltranspeptidase. The reaction was optimized for γ-glutamyl-p-nitroanilide. Key factors such as substrate specificity, pH, temperature, and the substrate mole ratio were all investigated. Kinetic studies of the acyl transfer reaction were described and the Hammett plot was constructed. This study indicated that the ratelimiting acylation reaction of γ-glutamyltranspeptidase can apparently be accelerated by either the electron-withdrawing or electron-donating substituents of γ-glutamyl anilides. The reaction could be catalyzed by the general acid and carboxy of Asp-433 or phenolic hydroxyl Tyr-444 may be the acid by autodock simulation for all prepared γ-glutamyl anilides.
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KEYWORD
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Escherichia coli γ-glutamyltranspeptidase, γ-glutamyl anilide analogues, kinetic studies, theanine
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