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[1]张飞龙,杨卫华,陈明亮,等.固定化天冬氨酸酶制备(R)-3-氨基丁酸[J].生物加工过程,2020,18(05):556-560.[doi:10.3969/j.issn.1672-3678.2020.05.002]
 ZHANG Feilong,YANG Weihua,CHEN Mingliang,et al.Preparation of(R)-3-aminobutyric acid by immobilized aspartase[J].Chinese Journal of Bioprocess Engineering,2020,18(05):556-560.[doi:10.3969/j.issn.1672-3678.2020.05.002]
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固定化天冬氨酸酶制备(R)-3-氨基丁酸()
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《生物加工过程》[ISSN:1672-3678/CN:32-1706/Q]

卷:
18
期数:
2020年05期
页码:
556-560
栏目:
出版日期:
2020-10-30

文章信息/Info

Title:
Preparation of(R)-3-aminobutyric acid by immobilized aspartase
文章编号:
1672-3678(2020)05-0556-05
作者:
张飞龙1杨卫华1陈明亮1严燕兵1唐云平2肖延铭1
1.长兴制药股份有限公司 工业生物催化与转化浙江省工程研究中心,浙江 长兴 313100; 2.浙江海洋大学 食品与医药学院,浙江 舟山 316022
Author(s):
ZHANG Feilong1YANG Weihua1CHEN Mingliang1YAN Yanbing1TANG Yunping2XIAO Yanming1
1.Zhejiang Engineering Research Center of Industrial Biocatalysis and Transformation,Changxing Pharmaceutical Co.,Ltd., Changxing,313100,China; 2.School of Food and Pharmacy,Zhejiang Ocean University,Zhoushan 316022,China
关键词:
天冬氨酸酶 巴豆酸(R)-3-氨基丁酸 固定化
分类号:
Q814.2
DOI:
10.3969/j.issn.1672-3678.2020.05.002
文献标志码:
A
摘要:
从芽孢杆菌Bacillus sp.YM55-1基因组中克隆得到天冬氨酸酶基因,以pET-28a(+)为载体构建天冬氨酸酶基因的表达载体pET-28a(+)-Asp,将天冬氨酸酶基因进行定点突变,在E.coli BL21(DE3)系统中实现了天冬氨酸酶的异源表达。利用重组的天冬氨酸酶,以氨水、(NH4)2SO4为辅料,将底物巴豆酸转化为(R)-3-氨基丁酸。将天冬氨酸酶的工程菌制备成固定化细胞,通过反应条件的优化研究,提高底物的转化率。结果表明:天冬氨酸酶最适pH为9.0,最适反应温度为40 ℃。在此反应条件下,加入30 g/L固定化细胞,转化22 h,(R)-3-氨基丁酸质量浓度达到220 g/L,对映体过量值e.e.s≥99.95%,底物转化率达到98%,固定化细胞重复使用次数不低于24次。
Abstract:
Aspartate ammonia-lyase gene of Bacillus sp.YM55-1 was cloned into the expression vector pET-28a(+).By site-directed mutagenesis of aspartate ammonia-lyase gene,the heterologous expression was obtained in E. coli BL21(DE3)cells.Conversion of crotonic acid to(R)-3-aminobutyric acid was performed by the recombinant aspartate ammonia-lyase in the presence of ammonia and ammonium sulfate.The aspartate ammonia-lyase engineering bacteria were prepared into immobilized cells,and then enzymatic reaction conditions were optimized to achieve higher production of(R)-3-aminobutyric acid.The optimum pH and reaction temperature was 9.0 and 40 ℃,respectively.Under these conditions,immobilized cells of 30 g/L were added and incubated for 22 h.Finally,the production of(R)-3-aminobutyric acid was 220 g/L,enantiomeric excess was more than 99.95%,conversion up to 98% and immobilized cells could be reused no less than 24 times.

参考文献/References:

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备注/Memo

备注/Memo:
收稿日期:2018-09-13修回日期:2020-03-24
基金项目:国家高技术研究发展计划(863计划)(2015AA021004)
作者简介:张飞龙(1989—),男,安徽阜阳人,助理工程师,研究方向:生物化学与分子生物学,E-mail:zhangfl379@163.com; 肖延铭(联系人),工程师,E-mail:hrbxym1286@yeah.net
引文格式:张飞龙,杨卫华,陈明亮,等.固定化天冬氨酸酶制备(R)-3-氨基丁酸[J].生物加工过程,2020,18(5):556-560.
ZHANG Feilong,YANG Weihua,CHEN Mingliang,et al.Preparation of(R)-3-aminobutyric acid by immobilized aspartase[J].Chin J Bioprocess Eng,2020,18(5):556-560..
更新日期/Last Update: 2020-09-30