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投稿时间：2022-12-28
投稿时间：2022-12-28
中文摘要: γ-氨基丁酸（γ-aminobutyric acid，GABA）是一种非蛋白天然氨基酸，具有多种生物活性。谷氨酸脱羧酶（glutamate decarboxylase，GAD）能将L-谷氨酸或谷氨酸钠不可逆地催化为γ-氨基丁酸。为探究GAD 酶学性质，从植物乳杆菌CPRGJ（Lactobacillus plantarum）中克隆表达GAD，利用镍柱亲和层析对重组GAD 进行纯化并探究其酶学性质。结果表明，GAD 的酶学性质为最适温度45 ℃、最适pH5.0、辅酶5-磷酸吡哆醛摩尔浓度250μmol/L，此时有最大酶活力；有机试剂中正丁醇对GAD 有强抑制作用，通过Lineweaver Burk 方程可得到GAD 的米氏常数Km为19.988 mmol/L，Vmax 为0.263 mmol/（L·min）。
Abstract:Gamma-aminobutyric acid（GABA）is a natural，non-protein amino acid with a variety of biological activities. Glutamate decarboxylase（GAD）can irreversibly catalyze L-glutamate or sodium glutamate into GABA.To investigate the enzymatic properties of GAD，it was cloned and expressed from Lactobacillus plantarum CPRGJ，and the recombinant GAD was purified using nickel column affinity chromatography. Its enzymatic properties were then explored.The results showed that the enzymatic properties of GAD were as follows：the optimum temperature was 45 ℃，the optimum pH was 5.0，and the molar concentration of the coenzyme pyridoxal 5′-phosphate（PLP）was 250 μmol/L，at which the enzyme exhibited maximum activity. Among organic reagents，N-butanol had a strong inhibitory effect on GAD.The Michaelis constant（Km）of GAD was determined using the Lineweaver-Burk equation，yielding a value of 19.988 mmol/L，and the maximum reaction velocity（Vmax）was 0.263 mmol/（L·min）.
keywords: γ-aminobutyric acid glutamate decarboxylase cloning enzymatic properties Lactobacillus plantarum
文章编号：202421025 中图分类号： 文献标志码：
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