Spitsberg V L, Gorewit R C
Department of Animal Science, Cornell University, Ithaca, New York, 14853, USA.
Protein Expr Purif. 1998 Jul;13(2):229-34. doi: 10.1006/prep.1998.0901.
Bovine milk xanthine oxidase (XO) was isolated and purified from milk fat globule membrane (MFGM). The method included the following steps: solubilization of XO from MFGM in 200 mM dithiothreitol (DTT) at pH 8.0, fractionation of solubilized proteins with ammonium sulfate, chromatography on DEAE-Sepharose with gradient elution, and rechromatography of the XO fraction for final purification. The method is highly reproducible, is comparatively simple, and provides highly pure enzyme. Purified XO, analyzed by (8%) SDS-PAGE, had only one band of 140-150 kDa. XO showed a high specific activity of 2.5 units/mg of protein and an A280: A450 ratio of 4.8.
从乳脂肪球膜（MFGM）中分离并纯化了牛乳黄嘌呤氧化酶（XO）。该方法包括以下步骤：在pH 8.0的200 mM二硫苏糖醇（DTT）中从MFGM溶解XO，用硫酸铵分级分离溶解的蛋白质，在DEAE-琼脂糖上进行梯度洗脱色谱，以及对XO级分进行再色谱以进行最终纯化。该方法具有高度可重复性，相对简单，并能提供高纯度的酶。通过（8%）SDS-PAGE分析，纯化的XO只有一条140 - 150 kDa的条带。XO显示出2.5单位/毫克蛋白质的高比活性和4.8的A280:A450比值。
