| 林谦，姜康怡，韦素娟，郭丹妮，农石蓉，高敏.产GABA发酵乳杆菌的筛选、发酵条件优化
及其谷氨酸脱羧酶基因的克隆[J].广东农业科学,2014,41(8):192-197 |
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产GABA发酵乳杆菌的筛选、发酵条件优化
及其谷氨酸脱羧酶基因的克隆 |
| Screening of GABA-producing Lactobacillus fermentum,optimized fermentation for GABA production andcloning of its glutamate decarboxylase gene |
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| DOI： |
| 中文关键词: 发酵乳杆菌 酌-氨基丁酸 谷氨酸脱羧酶 克隆 序列分析 |
| 英文关键词: Lactobacillus fermentum 酌-Aminobutyric acid Glutamate decarboxylase cloning sequence analysis |
| 基金项目:广西自然科学基金(2011GXNSFB018046）；广西
高校特色专业及课程一体化建设项目(GXTSZY220，GXTSZY022） |
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| 中文摘要: |
| 从酸菜汁中分离到一株具有谷氨酸脱羧酶活力的乳酸菌YS2，经过16S rDNA 序列分析鉴定为发酵乳
杆菌(Lactobacillus fermentum）。YS2 在含1% L-谷氨酸钠的MRS 培养基中，GABA 的产量达到4.37 g/L。对YS2 产
GABA 的碳源、氮源、初始pH 进行了初步优化，确定最佳条件为院以蔗糖为主要碳源，以蛋白胨和牛肉膏为氮源，初
始pH 6.0，在此条件下，GABA 产量达到5.68 g/L。通过PCR 顺利地扩增出了YS2 菌株的谷氨酸脱羧酶gadB 基因，将
PCR 产物与表达载体pEASY-E1 连接。测序结果表明，YS2 的谷氨酸脱羧酶与Lactobacillus fermentum F-6 的谷氨酸
脱羧酶序列一致性达到99%。重组酶与组氨酸标签融合表达，经镍柱亲和层析纯化出融合蛋白，电泳显示出56 kDa
的目的条带，纯化的重组酶表现出了酶活性(1.06 U/mg）。 |
| 英文摘要: |
| A strain of lactic acid bacteria, YS2, capable of production of aminobutyric acid (GABA), was isolated
from pickled vegetable juice, and was identified as Lactobacillus fermentum by 16S rDNA sequence analysis. When YS2
was grown in MRS medium containing 1% L-monosodium glutamate (MSG), GABA yield reached 4.37 g/L. The optimal
growth condition for GABA production was determined as follows: sucrose as carbon source, peptone plus beef extract as
nitrogen source, initial pH at 6.0. Under the condition, GABA yield reached 5.68 g/L. The gene encoding glutamate
decarboxylase, gadB, was amplified by PCR, and inserted into pEASY-E1, an expression vector for E. coli. Sequence
analysis showed that the GAD from YS2 had 99% identity with that of Lactobacillus fermentum F-6. GAD was expressed
with N-terminal hexahistidine tag fusion, and SDS-PAGE showed a major band of approximatela 56 kDa after purification
by Ni-NTA affinity chromatography. The specific activity of purified GAD reached 1.06 U/mg. |
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