鸭肝丁酰胆碱酯酶的纯化与酶学性质研究  

Purification and characterization of the butyrylcholinesterase from duck liver

作　　者：朱鸿[1] 李想韵[1] 邓玉[1] 王松[1] 付伟丽[1] 唐靓婷[1] 诰赵伟[1] 唐云明[1] 

机构地区：[1]西南大学生命科学学院,三峡库区生态环境教育部重点实验室,重庆市甘薯工程研究中心,重庆400715

出　　处：《食品工业科技》2011年第1期95-99,共5页Science and Technology of Food Industry

基　　金：重庆市科委资助项目(CSCT,2004AC1012)

摘　　要：目的:获得鸭肝丁酰胆碱酯酶纯品并对其酶学性质进行研究。方法:采用丙酮脱脂、酸沉淀、硫酸铵分级沉淀、DEAE-Sepharose阴离子交换层析和Sephacryl S-200凝胶层析方法,分离纯化鸭肝丁酰胆碱酯酶;采用SDS-聚丙烯酰胺凝胶电泳法进行纯度鉴定和酶相对分子量测定。结果:从鸭肝中分离纯化获得电泳纯的丁酰胆碱酯酶,纯化倍数为156.45倍,酶活回收率为23.60%,比活达17.21U/mg。酶相对分子量为388.85kDa,亚基相对分子量为64.70kDa。推测该酶由六个相同亚基构成。该酶最大紫外吸收为278nm。酶催化碘化硫代丁酰胆碱水解的最适pH为8.0,最适温度为35℃。该酶在pH3.0～10.0区域较稳定;在40℃以下处理1h,酶活力保持稳定。Zn2+、Mn2+和Cu2+对该酶具有显著的抑制作用。以碘化丁酰硫代胆碱为底物,测定该酶的表观Km为71.15μmol/L,没有过量底物抑制现象。结论:成功分离纯化获得丁酰胆碱酯酶,该酶具有较好的酸碱耐受性。Objective：To obtain butyrylcholinesterase from duck liver and study on the characterization of the purified product.Methods： The butyrylcholinesterase was extracted by acetone treatment, acid precipitation, ammonium sulfate precipitation, and ion-exchange chromatography on DEAE-Sepharose and gel filtration on Sephacryl S-200. SDS-PAGE was used to identify the purity and relative molecular of the butyrylcholinesterase.Results：The enzyme was purified to electrophoretic homogeneity. It was purified 156.45-fold and the activity recovery 23.60% was obtained.Its specific activity was 17.21 U/mg.The relative molecular weight of this butyrylcholinesterase was 388.85kDa,and the weight of subunit was 64.70kDa.The butyrylcholinesterase consisted of six identical subunits. Ultraviolet spectrum showed a maximum absorption at 278nm.The optimum pH and temperature of the enzyme for the hydrolysis of S-Butyrylthiocholine iodide were 8.0 and 35%, respectively.The enzyme was stable in the pH ranges of 3.0-10.0 under 35% and at temperatures below 45℃.Zn2＋ , Mn2＋ ,Cu2＋ had significant inhibition on this enzyme and the enzyme could not be inhibited by excess substrate. The apparent Km of this enzyme was 71.15μmol/L at pH8.0 and 35℃. Conclusion： The butyrylcholinesterase was successfully purified, and this butyrylcholinesterase showed good acid-base tolerance.

关 键 词：鸭肝 丁酰胆碱酯酶 分离纯化 性质 

分 类 号：TS201.1[轻工技术与工程—食品科学]