THE RIBONUCLEASE-ACTIVITY OF NUCLEOLAR PROTEIN B23
THE RIBONUCLEASE-ACTIVITY OF NUCLEOLAR PROTEIN B23
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DOI：
10.1093/nar/23.19.3974
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发表时间：
1995-10-11
影响因子：
14.9
通讯作者：
OLSON, MOJ
中科院分区：
文献类型：
--
作者：
HERRERA, JE;SAVKUR, R;OLSON, MOJ
蛋白B23是一种丰富的核仁蛋白和假定的核糖体组装因子，用rna包埋凝胶和高氯酸沉淀法分析了该蛋白的核糖核酸酶活性。该蛋白的三种纯化细菌表达形式B23.1、B23.2和标记为B23.1的n端多组氨酸以及天然蛋白被发现具有核糖核酸酶活性。重组B23.1比重组B23.2的特异性活性高5倍以上。该活性对人胎盘核糖核酸酶抑制剂不敏感，但对小牛胸腺DNA呈剂量依赖性抑制。该酶在很宽的pH范围内表现出活性，pH值为7.5时明显达到最佳。低浓度的Ca2+、Mg2+或NaCl的存在可刺激其活性，但不依赖于此。Ca2+的作用是饱和的，在性质上只是刺激。相比之下，Mg2+和NaCl表现出最佳的刺激浓度，并且在此浓度以上均抑制核糖核酸酶。这些数据表明蛋白质B23具有内在的核糖核酸酶活性，蛋白质B23在核核体含有核糖体RNA的亚室中的位置表明其核糖核酸酶活性在核糖体RNA的加工中起作用。
Protein B23 is an abundant nucleolar protein and putative ribosome assembly factor, The protein was analyzed for ribonuclease activity using RNA-embedded gels and perchloric acid precipitation assays. Three purified bacterially expressed forms of the protein, B23.1, B23.2 and an N-terminal polyhistidine tagged B23.1 as well as the natural protein were found to have ribonuclease activity. However, the specific activity of recombinant B23.1 was similar to 5-fold greater than that of recombinant B23.2. The activity was insensitive to human placental ribonuclease inhibitor, but was inhibited by calf thymus DNA in a dose dependent manner. The enzyme exhibited activity over a broad range of pH with an apparent optimum at pH 7.5. The activity was stimulated by but not dependent on the presence of low concentrations of Ca2+, Mg2+ or NaCl. The Ca2+ effect was saturable and only stimulatory in nature. In contrast, Mg2+ and NaCl exhibited optimal concentrations for stimulation and both inhibited the ribonuclease at concentrations above these optima, These data suggest that protein B23 has intrinsic ribonuclease activity, The location of protein B23 in subcompartments of the nucleolus that contain preribosomal RNA suggests that its ribonuclease activity plays a role in the processing of preribosomal RNA.