Acetylcholinesterase （ACHE） plays an important role in enzyme-based detection of pesticides in the environment. In this paper, ACHE from the Triton X-100 extract of brain tissues of Oreochromis aurea was purified by...Acetylcholinesterase （ACHE） plays an important role in enzyme-based detection of pesticides in the environment. In this paper, ACHE from the Triton X-100 extract of brain tissues of Oreochromis aurea was purified by （NH4）2SO4 fractional precipitation, Sephadex G-100 gel filtration, and DEAE-cellulose ion exchange chromatography. Certain biochemical characterizations of the purified enzyme and inhibition of pesticides on the enzyme were also studied. The specific activity of this purified enzyme was 20.628 U/mg protein, fold of purification was 139, and recovery was 22.1%. The optimal temperature of this enzyme was between 35-40 ℃, and optimal pH was between 7.5-8.0. The Michaelis constant （Kin） for acetylthiocholine iodide was 0.183 mmol/L. The enzyme activity was inhibited by excess substrate, and optimal substrate concentration was 6 mmol/L. Four pesticides （dichlorvos, phoxim, triazophos, and methomyl） exhibited strong inhibitions on this enzyme with IC50 less than 5 μg/mL. This study suggests that Oreochromis aurea （tilapia） could be a good enzyme source for pesticide monitoring in water environments.展开更多
基金supported by Huizhou Science and Technology Planning Project Foundation (Grant No.2008P60)
文摘Acetylcholinesterase （ACHE） plays an important role in enzyme-based detection of pesticides in the environment. In this paper, ACHE from the Triton X-100 extract of brain tissues of Oreochromis aurea was purified by （NH4）2SO4 fractional precipitation, Sephadex G-100 gel filtration, and DEAE-cellulose ion exchange chromatography. Certain biochemical characterizations of the purified enzyme and inhibition of pesticides on the enzyme were also studied. The specific activity of this purified enzyme was 20.628 U/mg protein, fold of purification was 139, and recovery was 22.1%. The optimal temperature of this enzyme was between 35-40 ℃, and optimal pH was between 7.5-8.0. The Michaelis constant （Kin） for acetylthiocholine iodide was 0.183 mmol/L. The enzyme activity was inhibited by excess substrate, and optimal substrate concentration was 6 mmol/L. Four pesticides （dichlorvos, phoxim, triazophos, and methomyl） exhibited strong inhibitions on this enzyme with IC50 less than 5 μg/mL. This study suggests that Oreochromis aurea （tilapia） could be a good enzyme source for pesticide monitoring in water environments.
