牛肝谷氨酸脱氢酶的分离纯化及部分酶学性质

doi:10.7506/spkx1002-6630-201513033

食品科学

牛肝谷氨酸脱氢酶的分离纯化及部分酶学性质

王丹，傅婷，万骥，唐云明*

西南大学生命科学学院，重庆市甘薯工程研究中心，三峡库区生态环境教育部重点实验室，重庆 400715

出版日期:2015-07-15 发布日期:2015-07-08

Isolation, Purification and Partial Characterization of Glutamate Dehydrogenase from Bovine Liver

WANG Dan, FU Ting, WAN Ji, TANG Yunming*

Key Laboratory of Eco-environments in Three Gorges Reservoir Region, Ministry of Education, Chongqing Sweetpotato Engineering
Research Center, School of Life Science, Southwest University, Chongqing 400715, China

Online:2015-07-15 Published:2015-07-08

摘要/Abstract

摘要：

取新鲜牛肝，经匀浆、缓冲液抽提、正丁醇脱脂、硫酸铵分级沉淀、DEAE-Sepharose离子交换层析、Superdex-200凝胶过滤层析等方法纯化，获得了电泳纯的牛肝谷氨酸脱氢酶（glutamate dehydrogenase，GDH）。经过纯化后的结果：GDH的酶比活力为306.06 U/mg，纯化倍数为93.60 倍，酶活回收率为23.12%。GDH的分子质量为380.2 kD，亚基分子质量约为61.7 kD。酶学性质研究结果表明：GDH的最适反应温度为50 ℃，在温度为30 ℃以下时较稳定；最适反应pH值为8.2，该酶对烟酰胺腺嘌呤二核苷酸（nicotinamide adenine dinucleotide，NADH）的Km值为0.696 mmol/L。甲醇、乙醇、异丙醇、Cd2+、Co2+、Ca2+、Ag+、Pb2+、Zn2+、Cu2+对该酶具有抑制作用，低浓度Ba2+、Mg2+、K+、Li+与乙二胺四乙酸（ethylenediamine tetraacetic acid，EDTA）对其具有一定的激活作用。

关键词: 牛肝, 谷氨酸脱氢酶, 分离纯化, 酶学性质

Abstract:

Electrophoresis-purity glutamate dehydrogenase (GDH) from bovine liver was obtained after homogenization,
buffer solution extraction, n-butanol degreasing, ammonium sulfate fractional precipitation, DEAE-Sepharose ion exchange
chromatography and Superdex-200 gel filtration. After purification, the specific activity of GDH was 306.06 U/mg, with
93.60-fold purification; meanwhile, the recovery rate was 23.12%. The molecular weight of GDH was approximately
380.2 kD, in which the subunit molecular weight was roughly 61.7 kD. The results of enzymatic characterization illustrated
that the optimal reaction temperature for GDH was 50 ℃, and it was relatively stable at a temperature below 30 ℃. The
optimal reaction was pH 8.2, and its Km was 0.696 mmol/L towards NADH. The enzyme activity of GDH was inhibited
by methanol, ethanol, isopropanol and metal ions such as Cd2+, Co2+, Ca2+, Ag+, Pb2+, Zn2+, and Cu2+, but enhanced to some
extent by low concentrations of Ba2+, Mg2+, K+, Li+ and EDTA.

Key words: bovine liver, glutamate dehydrogenase, isolation and purification, enzymatic properties

中图分类号:

Q946.5

王丹，傅婷，万骥，唐云明*. 牛肝谷氨酸脱氢酶的分离纯化及部分酶学性质[J]. 食品科学, doi: 10.7506/spkx1002-6630-201513033.

WANG Dan, FU Ting, WAN Ji, TANG Yunming*. Isolation, Purification and Partial Characterization of Glutamate Dehydrogenase from Bovine Liver[J]. FOOD SCIENCE, doi: 10.7506/spkx1002-6630-201513033.

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牛肝谷氨酸脱氢酶的分离纯化及部分酶学性质

Isolation, Purification and Partial Characterization of Glutamate Dehydrogenase from Bovine Liver

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