The purification of a nuclease from rat-liver mitochondria is described. The mitochondria are rendered soluble by treatment with Triton X-100 and, after fractionation with ammonium sulphate and acetone, the active fraction is further purified by chromatography on DEAE-cellulose and Sephadex G-75 to give a purification of over 700-fold. 2. The purified enzyme was only very slightly contaminated with deoxyribonuclease II, phosphodiesterase and phosphomonoesterase. The individual activities of these enzymes did not exceed 0.1% of the activity of the liver nuclease. 3. The purified enzyme attacked RNA more rapidly than denatured DNA and hydrolysed native DNA more slowly than denatured DNA. 4. There is some evidence to suggest that the nucleolytic activity of the purified preparation towards native DNA, denatured DNA and RNA is associated with a single protein. 5. The enzyme is relatively labile but is stabilized in the presence of 20% (w/v) glycerol or 10mm-2-mercaptoethanol.
摘要
本文描述了从大鼠肝脏线粒体中纯化一种核酸酶的方法。用 Triton X - 100 处理使线粒体可溶，经硫酸铵和丙酮分级分离后，活性部分再通过 DEAE - 纤维素和 Sephadex G - 75 柱层析进一步纯化，纯化倍数超过 700 倍。2. 纯化后的酶仅轻微污染有脱氧核糖核酸酶 II、磷酸二酯酶和磷酸单酯酶。这些酶的各自活性不超过肝脏核酸酶活性的 0.1%。3. 纯化后的酶对 RNA 的作用比对变性 DNA 更快，对天然 DNA 的水解比对变性 DNA 更慢。4. 有一些证据表明，纯化制剂对天然 DNA、变性 DNA 和 RNA 的核酸分解活性与单一蛋白质有关。5. 该酶相对不稳定，但在 20%（w/v）甘油或 10 mM 2 - 巯基乙醇存在下会稳定。
