产谷氨酸脱羧酶重组枯草芽孢杆菌的构建与食品级表达

Construction of A Recombinant Bacillus subtilis and Food-Grade Expression of Glutamate Decarboxylase

DOI：10.3969/j.issn.1673-1689.2020.02.004

中文关键词: 谷氨酸脱羧酶 γ-氨基丁酸 食品级表达

英文关键词: glutamate decarboxylase,γ-aminobutyric acid,food-grade expression

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中文摘要:

谷氨酸脱羧酶（GAD）是生物合成γ-氨基丁酸（GABA）的关键酶。本研究构建了4株无抗标记的重组枯草芽孢杆菌，首次实现了乳酸乳球菌（Lactococcus lactisssp. lactis IL 1403）来源的谷氨酸脱羧酶基因在枯草芽孢杆菌中的食品级表达。通过比较4株重组枯草芽孢杆菌的生长曲线和发酵酶活曲线，筛选出产酶效率最高的重组菌株B.subtilisWB600/pUB-P43-gadB(opt)-dal，该重组菌在初始发酵培养基中发酵42 h后发酵酶活可达4.1 U/mL。通过调整培养基成分，重组菌B.subtilisWB600/pUB-P43-gadB(opt)-dal的发酵酶活最高达到7.4 U/mL，与初始相比酶活提高了79%，是目前已报道重组枯草芽孢杆菌产谷氨酸脱羧酶酶活的最高水平。

英文摘要:

Glutamate decarboxylase（GAD）is a key enzyme that can be used to biosynthesize γ-aminobutyric acid（GABA）. In our study，four antibiotic-free marker recombinantBacillus subtilisstrains were constructed and glutamate decarboxylase derived fromLactococcus lactisssp. lactis IL 1403 was expressed in those strains. By comparing growth curves and GAD production curves of the four recombinant strains，the recombinant strainB.subtilisWB600/ pUB-P43-gadB（opt）-dal with highest enzyme activity was selected. After fermentation of the B.subtilis WB600/ pUB-P43-gadB（opt）-dal for 42 h in primary fermentation medium，the volumetric activity of the fermented broth reached 4.1 U/mL. By modifying of the composition of fermentation medium，volumetric activity of the fermented broth was increased by 79% and reached 7.4 U/mL，which was the highest activity of GAD for the reported recombinantBacillus subtilisstrains.

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