农药靶标乙酰胆碱酯酶的分离纯化及性质研究

Purification and characterization of acetylcholinesterase,a kind of pesticide target

采用了Sephadex G-25层析,DEAE-Sepharose Fast Flow离子交换层析和Sephacryl S-200凝胶过滤层析,对家蚕头部粗酶匀浆液中的乙酰胆碱酯酶进行纯化,得到的样品经聚丙烯酰胺凝胶电泳(PAGE)和SDS-PAGE检测均为一条带,用SDS-PAGE法测得其分子量为77.8kDa.酶的最适反应温度为37℃,最适pH为7.5,对底物碘化硫代乙酰胆碱(ATChI)的Km值为0.392 mmol/L,最适底物浓度为1.6 mmol/L,在1.6 mmol/L底物浓度以上观察到了底物抑制现象.

Acetylcholinesterase（AChE） of Bornbyx mori Linaeus, separated from the crude extract, was purified to electrophoretic homogeneity by Sephadex G-25 chromatography, DEAE-Sepharose Fast Flow ion-exchange chromatography and Sephacryl S-200 gel filtration, respectively. The Molecular weight of the purified enzyme was 77.8 kDa, measured by SDS-PAGE. The optimum temperature of the AChE was 37 ℃, above which the enzyme would be uristable. It exhibited optimum activity at pH 7.5. The Michaelis constant for acetylthiocholine iodide was 0. 392 mmol/L. The optimum concentration of substrate was 1.6 mmol/L, and the enzyme could be inhibitied by high level of acetvlthiocholine iodide.