POLIOVIRUS PROTEIN 2C CONTAINS 2 REGIONS INVOLVED IN RNA-BINDING ACTIVITY
POLIOVIRUS PROTEIN 2C CONTAINS 2 REGIONS INVOLVED IN RNA-BINDING ACTIVITY
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DOI：
10.1074/jbc.270.17.10105
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发表时间：
1995-04-28
影响因子：
4.8
通讯作者：
CARRASCO, L
中科院分区：
文献类型：
--
作者：
RODRIGUEZ, PL;CARRASCO, L
脊髓灰质炎病毒蛋白2C参与脊髓灰质炎病毒RNA复制，尽管2C的确切功能仍然未知。最近，显示当表达为与麦芽糖结合蛋白（MBP）的融合蛋白时，2C可以被纯化至高水平。预先提出的证据表明2C具有ATP酶和GTP酶活性;初步结果还表明2C与RNA相互作用（Rodriguez，P.L.，和Carrasco，L.（1993）J.Biol.Chem.268，8105-8110）。在本研究中，已经产生了2C的20种变体，并分析了它们的NTR和RNA结合活性。此外，描述了在因子Xa切割MBP（2）-2C融合蛋白后获得真正2C的简单方法。这项工作已经确定，2C有两个区域参与RNA结合：位于氨基酸21和45之间的NH 2-末端区域和位于氨基酸312和319之间的富含Arg的区域。NH 2-或COOH-末端RNA结合区的缺失消除了RNA结合。蛋白质2C的内部区域的缺失，包括核苷酸结合基序不影响RNA结合，而这种缺失破坏ATP酶和GT3酶活性。因此，NTR活性和蛋白2C的RNA结合能力位于分子的不同区域。
Poliovirus protein 2C is involved in poliovirus RNA replication, although the exact function of 2C is still unknown. Recently, it was shown that 2C can be purified to high levels when expressed as a fusion protein with maltose binding protein (MBP), Evidence was pre presented that 2C has ATPase and GTPase activities; preliminary results also indicated that 2C interacts with RNA (Rodriguez, P. L., and Carrasco, L. (1993) J. Biol. Chem. 268, 8105-8110), In the present study, 20 variants of 2C have been generated, and their NTPase and RNA binding activities were analyzed. Moreover, an easy procedure to obtain genuine 2C after factor Xa cleavage of an MBP(2)-2C fusion protein is described. This work has determined that 2C has two regions involved in RNA binding: a NH2-terminal region located between amino acids 21 and 45 and a COOH-terminal region involving an Arg-rich region located between amino acids 312 and 319. Deletion of either the NH2- or COOH-terminal RNA-binding region abolishes RNA binding. Deletion of an internal region of protein 2C that includes the nucleotide-binding motif does not affect RNA binding, whereas this deletion destroys ATPase and GTPase activities. Therefore, the NTPase activity and the RNA binding capacity of protein 2C are located in different regions of the molecule.