[1]郭旭舜,林裕添,贺敬东,等.大豆下胚轴谷丙转氨酶酶学性质分析[J].大豆科学,2021,40(01):82-88.[doi:10.11861/j.issn.1000-9841.2021.01.0082]
　GUO Xu-shun,LIN Yu-tian,HE Jing-dong,et al.Enzyme Properties Analysis of Glutamate-Pyruvate Transaminase in Soybean Hypocotyl[J].Soybean Science,2021,40(01):82-88.[doi:10.11861/j.issn.1000-9841.2021.01.0082]
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大豆下胚轴谷丙转氨酶酶学性质分析

《大豆科学》[ISSN:1000-9841/CN:23-1227/S] 卷: 第40卷 期数: 2021年01期 页码: 82-88 栏目: 出版日期: 2021-01-20
Title:
Enzyme Properties Analysis of Glutamate-Pyruvate Transaminase in Soybean Hypocotyl
文章编号:
2021,40(1)：082-088
作者:
郭旭舜; 林裕添; 贺敬东; 李昕昊; 张尚宏
(1.中山大学 生命科学学院，广东 广州 510275;2.中山大学 中山医学院，广东 广州 510080)
Author(s):
GUO Xu-shun;  LIN Yu-tian;  HE Jing-dong;  LI Xin-hao;  ZHANG Shang-hong
(1.School of Life Sciences, Sun Yat-sen University, Guangzhou 510275, China; 2.Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou 510080, China)
关键词:
大豆;  下胚轴;  谷丙转氨酶;  酶学性质;  液相色谱串联二级质谱;  茚三酮比色法
Keywords:
Glycine max;  Hypocotyl;  Glutamate-pyruvate transaminase;  Enzymatic properties;  LC-MS/MS;  Ninhydrin colorimetry
DOI:
10.11861/j.issn.1000-9841.2021.01.0082
文献标志码:
A
摘要:
大豆是一种重要的固氮农作物，在其种子萌发和初期幼苗生长过程中氨基酸代谢作用较强，表达活性较高的谷丙转氨酶（GPT）。为探明大豆下胚轴谷丙转氨酶的蛋白序列和酶学性质，以指导其在基因工程和农业生产中的应用，本研究通过对豆芽下胚轴蛋白组进行液相色谱串联二级质谱分析，确定大豆下胚轴所表达的GPT序列，利用谷氨酸和丙氨酸与茚三酮反应的显色差异测量GPT活力，并探究其酶学性质。结果显示：以茚三酮比色法测得大豆下胚轴谷丙转氨酶在35 ℃和pH7.0条件下对于丙酮酸的Km值为0.186 mmol?L-1，对于谷氨酸的Km值为1.05 mmol?L-1; Co2+、Mg2+和Ca2+对大豆谷丙转氨酶有激活效应，EDTA2-、尿素、Mn2+和Fe2+有抑制效应; 大豆下胚轴谷丙转氨酶具有活性的温度范围为10~60 ℃，pH范围为4.0~8.0，最适温度为35 ℃，最适pH7.0
Abstract:
Soybean (Glycine max) is an important nitrogen-fixing crop. It has strong amino acid metabolism and high expression of vigorous glutamate-pyruvate transaminase(GPT) during seed germination and early seedling growth. In this study, we identified the sequence of the GPT expressed in soybean hypocotyl through liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis, and described its kinetic characterization to guide its application in genetic engineeringand agricultural production. The Km values of this enzyme for glutamate and pyruvate were 1.05 and 0.186 mmol?L-1,respectively, measured by ninhydrin colorimetry based on the chromogenic difference between glutamate and alanine. Co2+, Zn2+, Ca2+ and Mg2+ had promotion effect, while EDTA2-, urea, Mn2+ and Fe2+ had inhibitory effect on the enzyme. The enzyme was active in the temperature range between 10 and 60 ℃, with the optimum temperature of 35 ℃, and active in the pH range between 4.0 and 8.0, with the optimum pH7.0, measured by separating glutamate and alanine through paper chromatography and calculating the enzyme activity.

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备注/Memo

中山大学本科教学质量工程项目（33000-18842502，33000-18832618）。

更新日期/Last Update: 2021-02-09