Lawton P, Sarciron E, Petavy A F
Laboratoire de Parasitologie, Faculté de Pharmacie, Université Claude-Bernard, Lyon, France.
J Parasitol. 1994 Oct;80(5):667-73.
The purification to homogeneity and the characterization of Echinococcus granulosus alkaline phosphatase (AP; EC 3.1.3.1) from hydatid cyst membranes are described. After n-butanol extraction, the parasite enzyme was sequentially purified by affinity chromatography on concanavalin A-sepharose followed by gel filtration. The purified protein (210 kDa) had a tetrameric structure composed of 4 56-kDa subunits. Its isoelectric point (4.8) and its kinetic parameters were determined (Km = 0.24 +/- 0.05 mmol/L; Vm = 173 +/- 21 nmol/min/mg protein for p-nitrophenylphosphate). The parasite enzyme differed from the host liver enzyme in its thermal stability, optimum reaction temperature, optimum pH, and catalytic parameters, but not in its apparent molecular weight. Furthermore, sera from patients infected with E. granulosus recognized the parasite AP on immunoblots, whereas uninfected controls were negative. These results as well as the role of this enzyme in the host-parasite relationship emphasize its potential importance as a diagnostic and prognostic antigen in the monitoring of hydatid infection.
本文描述了从细粒棘球绦虫包虫囊肿膜中纯化至同质的碱性磷酸酶（AP；EC 3.1.3.1）及其特性。经正丁醇提取后，寄生虫酶依次通过伴刀豆球蛋白A-琼脂糖亲和层析，随后进行凝胶过滤进行纯化。纯化后的蛋白质（210 kDa）具有由4个56 kDa亚基组成的四聚体结构。测定了其等电点（4.8）和动力学参数（对硝基苯磷酸酯的Km = 0.24±0.05 mmol/L；Vm = 173±21 nmol/min/mg蛋白质）。寄生虫酶在热稳定性、最佳反应温度、最佳pH值和催化参数方面与宿主肝脏酶不同，但表观分子量无差异。此外，细粒棘球绦虫感染患者的血清在免疫印迹中可识别寄生虫AP，而未感染的对照则为阴性。这些结果以及该酶在宿主-寄生虫关系中的作用强调了其作为包虫感染监测中诊断和预后抗原的潜在重要性。
