北京棒杆菌AS1.299高丝氨酸脱氢酶突变体D206G的酶学性质表征

doi:10.7506/spkx1002-6630-201307050

食品科学 ›› 2013, Vol. 34 ›› Issue (7): 240-244.doi: 10.7506/spkx1002-6630-201307050

北京棒杆菌AS1.299高丝氨酸脱氢酶突变体D206G的酶学性质表征

许金坤1,2，闵伟红1,2,*，詹冬玲1,2，方丽1,2，刘嘉1,2，申术霞1,2，郭永玲1,2

1.吉林农业大学食品科学与工程学院，吉林长春 130118；2.小麦和玉米深加工国家工程实验室，吉林长春 130118

收稿日期:2012-11-01 修回日期:2013-02-21 出版日期:2013-04-15 发布日期:2013-03-20
通讯作者: 闵伟红 E-mail:minwh2000@162.com

Characterization of Enzymatic Properties of High Homoserine Dehydrogenase-Producing Mutant D206G from Corynebacterium pekinense AS1.299

XU Jin-kun1,2，MIN Wei-hong1,2,*，ZHAN Dong-ling1,2，FANG Li1,2，LIU Jia1,2，SHEN Shu-xia1,2，GUO Yong-ling1,2

1. College of Food Science and Engineering, Jilin Agricultural University, Changchun 130118, China； 2. National Engineering Laboratory on Wheat and Corn Further Processing, Changchun 130118, China

Received:2012-11-01 Revised:2013-02-21 Online:2013-04-15 Published:2013-03-20
Contact: MIN Wei-hong E-mail:minwh2000@162.com

摘要/Abstract

摘要： 高丝氨酸脱氢酶(HSD)是天冬氨酸族氨基酸合成途径的关键酶，在苏氨酸和蛋氨酸生物合成中起到重要作用。将来自于北京棒杆菌AS1.299的HSD在E. coli BL21(DE3)中成功构建并进行高效异源表达。通过分子对接与同源序列比对发现，206位点参与底物结合，并且在家族中高度保守，因此对突变体D206G进行研究。结果表明：突变体最适pH7.5，最适反应温度37℃，半衰期为4h，并对金属离子和有机溶剂显示出良好的抗性。同野生型相比，突变体Km值减小，可能是由于侧链官能团减小，空间位阻降低，有利于底物结合。

关键词: 北京棒杆菌, 高丝氨酸脱氢酶, 定点突变, 酶学性质

Abstract: Homoserine dehydrogenase (HSD), a key enzyme involved in the biosynthesis of the aspartic acid family of amino acids, plays an important role in the synthesis of threonine and methionine. High HSD-producing mutant D206G from Corynebacterium pekinense AS1.299 was successfully constructed and heterologously expressed in E.coli BL21(DE3). Molecular docking and homologous alignment demonstrated that site 206 was a highly conservative sequence bound with substrate. The optimal pH, temperature and half-life of HSD from the mutant were 7.5, 37 ℃ and 4 h, respectively. In addition, it showed high resistance to metal ions and organic solvents. Compared with its wild-type counterpart, the Km of mutant D206G decreased, possibly due to a reduction in side-chain functional groups and consequent decrease in steric hindrance, thus promoting the enzyme-substrate binding.

Key words: Corynebacterium pekinense, homoserine dehydrogenase, site-directed mutagenesis, enzymatic

中图分类号:

Q814.9

许金坤1,2，闵伟红1,2,*，詹冬玲1,2，方丽1,2，刘嘉1,2，申术霞1,2，郭永玲1,2. 北京棒杆菌AS1.299高丝氨酸脱氢酶突变体D206G的酶学性质表征[J]. 食品科学, 2013, 34(7): 240-244.

XU Jin-kun1,2，MIN Wei-hong1,2,*，ZHAN Dong-ling1,2，FANG Li1,2，LIU Jia1,2，SHEN Shu-xia1,2，GUO Yong-ling1,2. Characterization of Enzymatic Properties of High Homoserine Dehydrogenase-Producing Mutant D206G from Corynebacterium pekinense AS1.299[J]. FOOD SCIENCE, 2013, 34(7): 240-244.

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北京棒杆菌AS1.299高丝氨酸脱氢酶突变体D206G的酶学性质表征

Characterization of Enzymatic Properties of High Homoserine Dehydrogenase-Producing Mutant D206G from Corynebacterium pekinense AS1.299

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