麦穗鱼脑乙酰胆碱酯酶(AChE)的纯化及其比较性研究

用PEG2000双水相萃取、DEAE-Sephadex A-50和Sephadex G-200方法分离纯化麦穗鱼Pseudorasbora parva脑中的乙酰胆碱酯酶(AChE),然后比较分析纯酶液与粗酶液中鱼脑AChE 的动力学特性和抗抑制性,以便更直接地了解底物与酶以及毒剂与酶的反应关系。研究结果表明,经过一系列步骤的纯化,最后所得的AChE是纯度较高的酶液;通过对AChE的动力学研究发现,纯化后的麦穗鱼脑AChE与底物之间的亲和力和其在粗酶状态时没有显著的差别,而且纯化后的鱼脑AChE对底物抑制作用更敏感;抗抑制性研究发现,纯化后的麦穗鱼脑AChE对经溴水氧化的马拉硫磷(malathion)和三唑磷(triazophos)的敏感性显著高于粗酶状态的AChE,酶更易受抑制。

Purification of Brain Acetylcholinerase (AChE) from Topmouth Gudgeon and Comparative Study between Crude and Purified AChE

Acetylcholinesterase (AChE) was purified from the brain tissues of topmouth gudgeon ( Pseudorasbora parva ) by PEG2000/phosphate salt two phases extraction, DEAE Sephadex A 50 and Sephadex G 200 chromatography. The comparative study of kinetic characters and resistance to inhibition by malaoxon and triazophos between crude and purified AChE from topmouth gudgeon was done. The result suggested that purified AChE is pure after the purification procedures; and the affinity of the brain AChE in purified state to substrate was no significantly different with that in crude state, while the inhibition of purified AChE to substrate was more sensitive than that of crude AChE; it was found in inhibition study that the sensitivity of purified AChE to both oxidized malathion and oxidized triazophos were significantly heigher than that of crude AChE.