Xia L, Chu K T, Ng T B
Department of Life Sciences, Shenzhen University, Shenzhen, China.
J Pept Res. 2005 Jul;66(1):1-8. doi: 10.1111/j.1399-3011.2005.00266.x.
A ribonuclease, with a molecular mass of 9 kDa and an N-terminal sequence resembling the sequence of a fragment of tRNA/rRNA cytosine-C5-methylase and a fragment of a alanyl-tRNA synthetase, was isolated from fresh fruiting bodies of the brown oyster mushroom Pleurotus ostreatus. The ribonuclease was purified using a very simple protocol that comprised ion-exchange chromatography on carboxymethyl (CM)-cellulose and affinity chromatography on Affi-gel blue gel. Subsequent gel filtration by fast protein liquid chromatography on Superdex 75 and sodium dodecyl sulfate (SDS) polyacrylamide gel electrophoresis revealed that the ribonuclease was purified after the first two chromatographic steps. The ribonuclease was adsorbed on CM-cellulose and Affi-gel blue gel. The ribonuclease exhibited the highest activity toward poly A, lower activity toward poly C, slight activity toward poly G, and indiscernible activity toward poly U. The enzyme was stimulated upon exposure to 1 microm Mg2+ and 10 microm Zn2+, but was inhibited by the following ions at 10 mm: Ca2+, Mg2+, Zn2+, Cu2+, Fe2+, Mn2+, and Fe3+. The ribonuclease required a pH of 8.0 and a temperature of 50-70 degrees C to express maximal activity. It had a Km of 60 microm toward yeast tRNA. It lacked mitogenic and HIV-1 reverse transcriptase inhibiting activities, but exerted antiproliferative activity toward leukemia L1210 cells.
从新鲜的糙皮侧耳子实体中分离出一种核糖核酸酶，其分子量为9 kDa，N端序列类似于tRNA/rRNA胞嘧啶-C5-甲基转移酶片段和丙氨酰-tRNA合成酶片段的序列。该核糖核酸酶采用非常简单的方案进行纯化，包括在羧甲基（CM）纤维素上进行离子交换色谱和在Affi-gel蓝凝胶上进行亲和色谱。随后通过Superdex 75快速蛋白质液相色谱进行凝胶过滤和十二烷基硫酸钠（SDS）聚丙烯酰胺凝胶电泳显示，在前两个色谱步骤后核糖核酸酶得到了纯化。该核糖核酸酶吸附在CM纤维素和Affi-gel蓝凝胶上。该核糖核酸酶对多聚A表现出最高活性，对多聚C活性较低，对多聚G有轻微活性，对多聚U活性不明显。该酶在暴露于1 μM Mg2+和10 μM Zn2+时受到刺激，但在10 mM的以下离子存在时受到抑制：Ca2+、Mg2+、Zn2+、Cu2+、Fe2+、Mn2+和Fe3+。该核糖核酸酶需要pH 8.0和50-70℃的温度来表达最大活性。它对酵母tRNA的Km为60 μM。它缺乏促有丝分裂和HIV-1逆转录酶抑制活性，但对白血病L1210细胞具有抗增殖活性。
