黄瓜过氧化物酶的分离纯化及酶学性质

doi:10.7506/spkx1002-6630-201411034

食品科学

黄瓜过氧化物酶的分离纯化及酶学性质

胡瑞斌，李星，王红杨，王洁，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:2014-06-15 发布日期:2014-07-03

Isolation, Purification and Characterization of Peroxidase from Cucumber

HU Rui-bin, LI Xing, WANG Hong-yang, WANG Jie, TANG Yun-ming

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

Online:2014-06-15 Published:2014-07-03

摘要/Abstract

摘要：

新鲜的黄瓜经匀浆、抽提、硫酸铵沉淀、CM-Sepharose 离子交换层析、Superdex-200凝胶过滤层析后获得电泳纯的过氧化物酶。该酶的比活力、回收率及纯化倍数分别为64 177.67 U/mg、9.58%、61.93。该酶的分子质量为41.15 kD，亚基分子质量为40.21 kD。该酶的最适温度和最适pH值分别为60 ℃和6。在25～45 ℃及pH 5～9的范围内非常的稳定。在测定条件下测得该酶的Km值为53.79 mmol/L。硫氰化钾对该酶活力基本无影响。尿素、K+、Mn2+、Ca2+、Mg2+、Ba2+、Cu2+对该酶都具有激活作用且浓度至50 mmol/L时酶活力分别被激活至112%、127%、113%、128%、139%、199%、348%，然而十二烷基磺酸钠、抗坏血酸和草酸对该酶有强烈的抑制作用；Zn2+、甲醇、乙醇和异丙醇对该酶有一定的抑制作用。

关键词: 黄瓜, 过氧化物酶, 分离纯化, 性质

Abstract:

Electrophoresis-purity peroxidase was extracted from fresh cucumber and purified through homogenization,
extraction, ammonium sulfate precipitation, CM-Sepharose and Superdex-200 chromatography. The specific activity,
recovery and purification fold of the peroxidase were 64 177.67 U/mg, 9.58% and 61.93, respectively. Molecular mass of
this purified enzyme was 40.21 kD as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE),
while native gel filtration confirmed a monomer form of 41.15 kD. Besides, the peroxidase, whose optimum temperature
and pH were 60 ℃ and 6, respectively, was comparatively stable in the temperature range of 25-45 ℃ and pH range of 5-9.
The purified peroxidase showed Km value of 53.79 mmol/L under definite conditions. In addition, its activity was scarcely
affected by potassium thiocyanate (KSCN). The peroxidase was found to be activated by urea, K+, Mn2+, Ca2+, Mg2+, Ba2+
and Cu2+ by 12%, 27%, 13%, 28%, 39%, 99% and 248% at 50 mmol/L, respectively. However, the peroxidase activity was
significantly inhibited by SDS, ascorbic acid and oxalic acid. Moreover, Zn2+, methanol, ethanol and isopropanol caused
partial inhibitory effects on the peroxidase.

Key words: cucumber, peroxidase, isolation and purification, characterization

胡瑞斌，李星，王红杨，王洁，唐云明. 黄瓜过氧化物酶的分离纯化及酶学性质[J]. 食品科学, doi: 10.7506/spkx1002-6630-201411034.

HU Rui-bin, LI Xing, WANG Hong-yang, WANG Jie, TANG Yun-ming. Isolation, Purification and Characterization of Peroxidase from Cucumber[J]. FOOD SCIENCE, doi: 10.7506/spkx1002-6630-201411034.

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黄瓜过氧化物酶的分离纯化及酶学性质

Isolation, Purification and Characterization of Peroxidase from Cucumber

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