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季也蒙毕赤酵母腈水解酶及其在3-羟基丙酸生物合成中的潜力。

Nitrile-hydrolyzing enzyme from Meyerozyma guilliermondii and its potential in biosynthesis of 3-hydroxypropionic acid.

作者信息

Zhang Qiang, Gong Jin-Song, Dong Ting-Ting, Liu Ting-Ting, Li Heng, Dou Wen-Fang, Lu Zhen-Ming, Shi Jin-Song, Xu Zheng-Hong

机构信息

School of Pharmaceutical Science, Jiangnan University, Wuxi, 214122, People's Republic of China.

National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, Wuxi, 214122, People's Republic of China.

出版信息

Bioprocess Biosyst Eng. 2017 Jun;40(6):901-910. doi: 10.1007/s00449-017-1754-6. Epub 2017 Mar 11.

Abstract

3-Hydroxypropionic acid (3-HP) is an important platform chemical in organic synthesis. Traditionally, 3-HP was produced by chemical methods and fermentation process. In this work, a novel enzymatic method was developed for green synthesis of 3-HP. A yeast strain harboring nitrile-hydrolyzing enzyme was newly isolated from environmental samples using 3-hydroxypropionitrile (3-HPN) as the sole nitrogen source. It was identified to be Meyerozyma guilliermondii CGMCC12935 by sequencing of the 18S ribosomal DNA and internal transcribed spacer, together with analysis of the morphology characteristics. The catalytic properties of M. guilliermondii CGMCC12935 resting cells were determined, and the optimum activity was achieved at 55 °C and pH 7.5. The enzyme showed broad substrate specificity towards nitriles, especially 3-HPN, aminoacetonitrile and 3-cyanopyridine. The presence of Ag, Pb and excess substrate inhibited the enzyme activity, whereas 5% (v/v) ethyl acetate had a positive effect on the enzyme activity. M. guilliermondii CGMCC12935 resting cells by addition of 3% glucose could thoroughly hydrolyze 500 mM 3-HPN into 3-HP within 100 h and the maximal accumulative production of 3-HP reached 216.33 mM, which was over twofolds than the control group with no additional glucose. And this work would lay the foundation for biological production of 3-HP in industry.

摘要

3-羟基丙酸(3-HP)是有机合成中一种重要的平台化合物。传统上,3-HP是通过化学方法和发酵工艺生产的。在这项工作中,开发了一种用于绿色合成3-HP的新型酶法。使用3-羟基丙腈(3-HPN)作为唯一氮源,从环境样品中新分离出一株含有腈水解酶的酵母菌株。通过对18S核糖体DNA和内部转录间隔区进行测序,并结合形态特征分析,将其鉴定为季也蒙毕赤酵母CGMCC12935。测定了季也蒙毕赤酵母CGMCC12935静息细胞的催化特性,其最佳活性在55°C和pH 7.5条件下实现。该酶对腈类具有广泛的底物特异性,尤其是对3-HPN、氨基乙腈和3-氰基吡啶。Ag、Pb和过量底物的存在会抑制酶活性,而5%(v/v)的乙酸乙酯对酶活性有积极影响。添加3%葡萄糖的季也蒙毕赤酵母CGMCC12935静息细胞能够在100小时内将500 mM的3-HPN完全水解为3-HP,3-HP的最大累积产量达到216.33 mM,比未添加葡萄糖的对照组高出两倍多。这项工作将为3-HP的工业化生物生产奠定基础。

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