乙醇对屎肠球菌全细胞谷氨酸脱羧酶表观活力的促进作用  

Ethanol improves the apparent activity of whole-cell glutamate decarboxylase in Enterococcus faecium

作　　者：杨胜远 徐小丽 梁永祥 YANG Shengyuan;XU Xiaoli;LIANG Yongxiang(College of Food Science and Engineering,Lingnan Normal University,Zhanjiang 524048,Guangdong,China)

机构地区：[1]岭南师范学院食品科学与工程学院,广东湛江524048

出　　处：《微生物学通报》2024年第3期758-768,共11页Microbiology China

基　　金：广东省重点建设学科科研能力提升项目(2022ZDJS079);广东省普通高校特色创新项目(2022KTSCX073);广东省本科高校教学质量与教学改革工程特色专业项目(粤教高函[2020]19号);岭南师范学院教学质量与教学改革工程特色专业项目(岭师教务[2020]228号)。

摘　　要：【背景】细胞内外物质的交换速率对全细胞转化活力具有重要影响。【目的】探讨乙醇对全细胞谷氨酸脱羧酶(glutamate decarboxylase,GAD)表观活力的影响,提高屎肠球菌全细胞转化法合成γ-氨基丁酸的产量。【方法】通过比较乙醇对GAD纯酶和全细胞GAD酶学特性、细胞结构、细胞膜通透性等的影响,探究乙醇对全细胞GAD活力的促进机制。【结果】低浓度乙醇对全细胞GAD活力有显著促进作用,最适浓度为7.5%;在该浓度下,全细胞GAD表观活力提升41.63%,乙醇对全细胞GAD最适反应pH无影响,但降低了全细胞GAD活力对胞外pH变化的敏感性,减小了米氏常数(Km);7.5%乙醇对GAD纯酶影响不显著(P>0.05),未造成GAD外泄和细胞破损,仅在与细胞共存的反应体系才具有促进作用。结果表明,7.5%乙醇可改变细胞的透性,提升细胞传质速率,从而促进全细胞GAD的表观活力。【结论】乙醇价格低廉、安全性高,对全细胞GAD的促进作用在GABA工业上具有很好的应用前景。[Background]The exchange rate of substances inside and outside cells has a significant effect on the transformation activity of whole-cells.[Objective]To increase the yield ofγ-aminobutyric acid(GABA)synthesized by whole-cell transformation,we investigated the improving effect of ethanol on the apparent activity of whole-cell glutamate decarboxylase(GAD)in Enterococcus faecium.[Methods]We explored the mechanism of ethanol in improving the apparent activity of whole-cell GAD by comparing the effects of ethanol on the enzymatic properties of pure GAD and whole-cell GAD,cell structure,and membrane permeability.[Results]Low-concentration ethanol significantly promoted the activity of whole-cell GAD with the optimal concentration of 7.5%.At this concentration,the apparent activity of whole-cell GAD was increased by 41.63%,and the optimal reaction pH of whole-cell GAD did not change.However,the sensitivity of whole-cell GAD to the changes of extracellular pH and the Michaelis constant(Km)of whole-cell GAD were both reduced.This concentration of ethanol had no significant effect on pure GAD(P>0.05),and did not cause GAD leakage or cell damage.In addition,7.5%ethanol increased the activity of GAD only when it coexisted with the cells in reaction system.The results indicated that 7.5%ethanol altered the cell permeability and enhanced the mass transfer rate of cells,thereby improving the apparent activity of whole-cell GAD.[Conclusion]Due to the low price and high safety,ethanol demonstrates great potential of application in the improvement of whole-cell GAD activity for the industrial production of GABA.

关 键 词：乙醇 谷氨酸脱羧酶 生物转化 Γ-氨基丁酸 促进作用 屎肠球菌 

分 类 号：TQ921.4[轻工技术与工程—发酵工程]