Ben Bacha Abir, Daihan Sooad K, Moubayed Nadine M S, Mejdoub Hafedh
Biochemistry Department, Science College, King Saud University, PO Box 22452, Riyadh 11495, Saudi Arabia.
Indian J Biochem Biophys. 2013 Jun;50(3):186-95.
A phospholipase A2 belonging to IIA group secretory PLA2 was isolated and purified to homogeneity from the intestine of common stingray (Dasyatis pastinaca) using acidic treatment (pH 1.5) and ammonium sulphate precipitation methods combined with single-column ion-exchange chromatography. The purified enzyme was found to be a glycosylated monomeric protein with a molecular mass of about 14 kDa. The stingray sPLA2-IIA had optimum activity at 45 degrees C, unlike known mammalian PLA2-IIAs, which show optimum activity at 37 degrees C. The purified enzyme exhibited a specific activity of 290 U/mg at optimal conditions (pH 9.5 and 45 degrees C) in the presence of 6 mM NaDC and 8 mM CaCl2 with egg yolk as substrate. The NH2-terminal sequence of the enzyme and some protein fragments obtained from its tryptic digestion were also determined. All sequences obtained were similar to those of sPLA2-IIA. The enzyme also showed good stability in the presence of organic solvents, acidic and alkaline pH media and high temperature conditions. Thus, the purified enzyme exhibited a number of unique and promising properties, making it a potential possible candidate for future applications in the treatment of phospholipid-rich industrial effluents and synthesis of useful preparations for the food production and processing industry.
利用酸性处理（pH 1.5）、硫酸铵沉淀法结合单柱离子交换色谱法，从黄貂鱼（Dasyatis pastinaca）的肠道中分离并纯化出一种属于IIA组分泌型磷脂酶A2的酶，使其达到同质状态。纯化后的酶是一种糖基化单体蛋白，分子量约为14 kDa。与已知在37℃表现出最佳活性的哺乳动物PLA2-IIA不同，黄貂鱼sPLA2-IIA在45℃具有最佳活性。在以蛋黄为底物、存在6 mM NaDC和8 mM CaCl2的最佳条件（pH 9.5和45℃）下，纯化后的酶表现出290 U/mg的比活性。还测定了该酶的NH2末端序列以及从其胰蛋白酶消化产物中获得的一些蛋白质片段。获得的所有序列均与sPLA2-IIA的序列相似。该酶在有机溶剂、酸性和碱性pH介质以及高温条件下也表现出良好的稳定性。因此，纯化后的酶表现出许多独特且有前景的特性，使其成为未来用于处理富含磷脂的工业废水以及合成食品生产和加工业有用制剂的潜在候选物。
