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中国生物工程杂志

China Biotechnology
China Biotechnology  2016, Vol. 36 Issue (5): 59-67    DOI: 10.13523/j.cb.20160509
    
Cloning and Expression of Dioxygenase Gene from Aeromonas sp. XJ-6 and Promoting Degradation of Tyr
YANG Yang1, YANG Jiang-ke2, XIONG Wei2, LIANG Jian-fang1, DUAN Wei-wei1, CHAO Qun-fang1
1. College of Life Science and Technology, Xinjiang University, Urumqi 830046, China;
2. School of Biology and Pharmaceutical Engineering, Wuhan Polytechnic University, Wuhan 430023, China
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Abstract  

Objective:To clone the dioxygenase dio6 gene from Aeromonas sp. XJ-6, and explore the function of dioxygenase dio6 to provide genetic resources for biodegradation of aromatic compounds. Methods:Dioxygenase dio6 gene from Aeromonas sp.XJ-6 genome was amplified by PCR and induciblly expressed in Escherichia coli BL21(DE3). The expression products of dio6 was purified by Ni-NTA.TLC and HPLC to detect the degradation effect of dioxygenase on Tyr, and the degradation products were also detected by LC-MS so as to analyze the possible degradation pathway. Results:The size of Aeromonas sp. XJ-6 dioxygenase gene dio6 was 1194bp, the expression product of dio6 purified by MCAC was 44.9kDa. HPLC analysis showed that dio6 has strong degradation ability for Tyr under the optimal conditions that the amount of enzyme is 60μl per reaction mixture, temperature 30℃.Metal ion Mg2+ and Ca2+ slightly inhibit the enzymatic reaction, and Mn2+、Zn2+、Cu2+、Fe2+ and Ca2+ can improve the substrate degradation, of which Mn2+ has the most impact on dioxygenase dio6. LC-MS analysis results showed that Tyr was turned into fumaric acid with the help of dioxygenase dio6. Conclusion:Aeromonas sp. XJ-6 dioxygenase dio6 is a kind of benzene ring-opening enzyme, which may be provided as a genetic resource for aromatic compounds biodegradation.



Key wordsPurification      Benzene ring      LC-MS      HPLC      Dioxygenas     
Received: 30 November 2015      Published: 04 January 2016
ZTFLH:  Q554+.7  
Cite this article:

YANG Yang, YANG Jiang-ke, XIONG Wei, LIANG Jian-fang, DUAN Wei-wei, CHAO Qun-fang. Cloning and Expression of Dioxygenase Gene from Aeromonas sp. XJ-6 and Promoting Degradation of Tyr. China Biotechnology, 2016, 36(5): 59-67.

URL:

https://manu60.magtech.com.cn/biotech/10.13523/j.cb.20160509     OR     https://manu60.magtech.com.cn/biotech/Y2016/V36/I5/59

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