The newly identified human nuclear protein NXP-2 possesses three distinct domains, the nuclear matrix-binding, RNA-binding, and coiled-coil domains
The newly identified human nuclear protein NXP-2 possesses three distinct domains, the nuclear matrix-binding, RNA-binding, and coiled-coil domains
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DOI：
10.1074/jbc.m201440200
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发表时间：
2002-06-07
影响因子：
4.8
通讯作者：
Osumi, T
中科院分区：
文献类型：
--
作者：
Kimura, Y;Sakai, F;Osumi, T
使用识别核基质蛋白的单克隆抗体，我们从人胎盘cDNA文库中选择了一个cDNA克隆。该cDNA编码939个氨基酸的蛋白质，命名为核基质蛋白NXP-2。北方印迹分析表明NXP-2在不同组织中有不同水平的表达。强制表达的绿色荧光蛋白标记的NXP-2以及内源性NXP-2定位于细胞核中并分布到核基质中。当在用于核基质制备的缓冲液中包含RNase A时，NXP-2从核基质中释放。使用NXP-2的绿色荧光蛋白标记的截短突变体进行功能结构域的作图。氨基酸326-353的区域负责核基质结合，并且包含与急性髓性白血病蛋白的核基质靶向信号相似的疏水氨基酸簇。通过Northwestern分析证明了中心区域（氨基酸500-591）是RNA结合所必需的，尽管NXP-2缺乏已知的RNA结合基序。预测氨基酸残基682-876的区域具有卷曲螺旋结构。RNA结合、核基质结合和卷曲螺旋结构域在结构上是分离的，这表明NXP-2在多种核功能中发挥重要作用，包括RNA代谢和核结构的维持。
Using a monoclonal antibody that recognizes a nuclear matrix protein, we selected a cDNA clone from a lambdagt11 human placenta cDNA library. This cDNA encoded a 939-amino acid protein designated nuclear matrix protein NXP-2. Northern blot analysis indicated that NXP-2 was expressed in various tissues at different levels. Forcibly expressed green fluorescent protein-tagged NXP-2 as well as endogenous NXP-2 was localized in the nucleus and distributed to the nuclear matrix. NXP-2 was released from the nuclear matrix when RNase A was included in the buffer for nuclear matrix preparation. Mapping of functional domains was carried out using green fluorescent protein-tagged truncated mutants of NXP-2. The region of amino acids 326-353 was responsible for nuclear matrix binding and contained a cluster of hydrophobic amino acids that was similar to the nuclear matrix targeting signal of acute myeloleukemia protein. The central region (amino acids 500-591) was demonstrated to be required for RNA binding by Northwestern analysis, although NXP-2 lacked a known RNA binding motif. The region of amino acid residues 682-876 was predicted to have a coiled-coil structure. The RNA-binding, nuclear matrix-binding, and coiled-coil domains are structurally separated, suggesting that NXP-2 plays important roles in diverse nuclear functions, including RNA metabolism and maintenance of nuclear architecture.