Halim H Y, Shaban E A, Hagag M M, Daoud E W, el-Asmar M F
Department of Biochemistry, Faculty of Medicine, Ain Shams University, Cairo, Egypt.
Toxicon. 1987;25(11):1199-207. doi: 10.1016/0041-0101(87)90138-3.
A venom exonuclease 'phosphodiesterase' (E.C. 3.1.4.1) has been purified from Cerastes cerastes venom by a combination of gel filtration on Sephadex G-100 superfine and ion exchange chromatography on DEAE-Sepharose. The enzyme showed a single band on PAGE and SDS-PAGE and had a molecular weight of 110,000. The final preparation was purified 28 fold. It had no carbohydrate and it did not have protease or 5'-nucleotidase activities. Optimum temperature for enzyme activity was 56 degrees C. The enzyme was rapidly inactivated when pre-incubated above 40 degrees C. Energy of activation (Ea) was calculated to be 0.913. The optimum pH was 9.0. Cysteine, glutathione, dithiothreitol, 2-mercaptoethanol, ADP and AMP inhibited the enzyme. Cysteine caused a non-competitive inhibition, while ADP showed a competitive inhibition. EDTA at a concentration of 0.5 mM caused complete inhibition of the enzyme, which could be reversed by the addition of Ca2+ or Mn2+.
已通过在Sephadex G - 100超细微粒上进行凝胶过滤和在DEAE - Sepharose上进行离子交换色谱相结合的方法，从角蝰毒液中纯化出一种毒液核酸外切酶“磷酸二酯酶”（E.C. 3.1.4.1）。该酶在聚丙烯酰胺凝胶电泳（PAGE）和十二烷基硫酸钠 - 聚丙烯酰胺凝胶电泳（SDS - PAGE）上显示为单一谱带，分子量为110,000。最终制剂纯化了28倍。它不含碳水化合物，也没有蛋白酶或5'-核苷酸酶活性。酶活性的最佳温度为56℃。当在40℃以上预孵育时，该酶会迅速失活。计算得出的活化能（Ea）为0.913。最佳pH值为9.0。半胱氨酸、谷胱甘肽、二硫苏糖醇、2 - 巯基乙醇、二磷酸腺苷（ADP）和一磷酸腺苷（AMP）会抑制该酶。半胱氨酸引起非竞争性抑制，而ADP表现出竞争性抑制。浓度为0.5 mM的乙二胺四乙酸（EDTA）会导致该酶完全抑制，添加钙离子（Ca2+）或锰离子（Mn2+）可使其逆转。
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