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http://dx.doi.org/10.4014/jmb.1405.05058

Cloning, Expression, and Characterization of Para-Aminobenzoic Acid (PABA) Synthase from Agaricus bisporus 02, a Thermotolerant Mushroom Strain  

Deng, Li-Xin (State Key Laboratory of Cellular Stress Biology, School of Life Science, Xiamen University)
Shen, Yue-Mao (Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Shandong University)
Song, Si-Yang (State Key Laboratory of Cellular Stress Biology, School of Life Science, Xiamen University)
Publication Information
Journal of Microbiology and Biotechnology / v.25, no.1, 2015 , pp. 66-73 More about this Journal
Abstract
The pabS gene of Agaricus bisporus 02 encoding a putative PABA synthase was cloned, and then the recombinant protein was expressed in Escherichia coli BL21 under the control of the T7 promoter. The enzyme with an N-terminal GST tag or His tag, designated GST-AbADCS or His-AbADCS, was purified with glutathione Sepharose 4B or Ni Sepharose 6 Fast Flow. The enzyme was an aminodeoxychorismate synthase, and it was necessary to add with an aminodeoxychorismate lyase for synthesizing PABA. AbADCS has maximum activity at a temperature of approximately 25℃ and pH 8.0. Magnesium or manganese ions were necessary for the enzymatic activity. The Michaelis-Menten constant for chorismate was 0.12 mM, and 2.55 mM for glutamine. H2O2 did distinct damage on the activity of the enzyme, which could be slightly recovered by Hsp20. Sulfydryl reagents could remarkably promote its activity, suggesting that cysteine residues are essential for catalytic function.
Keywords
Agaricus bisporus; para-aminobenzoic acid; aminodeoxychorismate synthase; aminodeoxychorismate lyase; Michaelis-Menten constant;
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