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| The Site-directed Mutation of Key Residues and the Analysis about Inhibitory Activity of Cassia obtusifolia Trypsin Inhibitor |
| XIANG Mian, ZHU Jian-quan, YU Ji-hua, LI Yang-yang, LI Juan-juan, LIU Zu-bi, WANG Wan-jun, LIAO Hai, ZHOU Jia-yu |
| Southwest Jiaotong University, Chengdu 610031, China |
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Abstract A trypsin inhibitor (CoTI1) from Cassia obtusifolia was attributed to the Kunitz-type trypsin inhibitor family. According to the sequence alignment, Arg86, Leu84 and Thr88 might be the key residues of CoTI1. In order to confirm the speculation, the three above residues were replaced as Asp by site-directed mutagenesis, respectively, and analysis the inhibitory activity of the mutants and CoT1 to trypsin and insects' digestive enzyme. Compared with CoT1, the inhibitory activity of the mutant CoTI1R86D to trypsin decreased most obviously, and the inhibitory effect of the original 93% was lost. CoTI1L84D lost 59% of the inhibitory effect; while the inhibitory activity of CoTI1T88D decreased by 64%. The average inhibitory activity decreased 88.7%, 57% and 60.7% to digestive enzymes of Helicoverpa armigera, Beet armyworm and Spodoptera litura, respectively. The result shows that Arg86, Leu84 and Thr88 are the key residues of CoT1, and it was useful for the molecular mechanism and anti-insects study of CoTI1.
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Received: 26 April 2016
Published: 25 October 2016
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Cite this article:
XIANG Mian, ZHU Jian-quan, YU Ji-hua, LI Yang-yang, LI Juan-juan, LIU Zu-bi, WANG Wan-jun, LIAO Hai, ZHOU Jia-yu. The Site-directed Mutation of Key Residues and the Analysis about Inhibitory Activity of Cassia obtusifolia Trypsin Inhibitor. China Biotechnology, 2016, 36(10): 15-20.
URL:
https://manu60.magtech.com.cn/biotech/10.13523/j.cb.20161003 OR https://manu60.magtech.com.cn/biotech/Y2016/V36/I10/15
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