...
  • 主题
高级搜索  >
历史搜索 清空历史
首页> 外文期刊>Biochemistry >Specific HIV-1 TAR RNA Loop Sequence and Functional Groups Are Required for Human Cyclin T1-Tat-TAR Ternary Complex Formation
【24h】

Specific HIV-1 TAR RNA Loop Sequence and Functional Groups Are Required for Human Cyclin T1-Tat-TAR Ternary Complex Formation

机译：人类细胞周期蛋白T1-Tat-TAR三元复合物的形成需要特定的HIV-1 TAR RNA环序列和功能基团。

获取原文
获取原文并翻译 | 示例
           
页面导航
  • 摘要
  • 著录项
  • 相似文献
  • 相关主题

摘要

Replication of human immunodeficiency virus requires Tat protein which activates elongation of RNA polymerase II transcription at the HIV-1 promoter through interaction with the cyclin T1 (CycT1) subunit of the positive transcription elongation factor complex (P-TEFvb). Tat binds directly through its transactivation domain to the CycTR1 suvbunit of the P-TEFbv and induces loop sequence specific binding of the P-TEFb onto nascent HIV-1 TAR RNA. By using a gel electrophoresis method and a comprehensive set of TAR loop mutants, we have identified the sequence and structural determinants for high-affinity CycT1-Tat-TAR ternary complex formation. Our results show that CycT1 and Tat binding to TAR RNA is highlycooperative, and a capacity of 85%, a Hill coefficient of 2.7, and a dissociation constant (K_D) of 2.45 nM were observed. These results indicate that there are three binding sites on TAR RNA. CycT1 does not bind TAR RNA in the absence of Tat, and Tat binding to TAR, while detectable, is very inefficient in the absence of CycT1. It is conceivable that the CycT1-Tat heterodimer directly binds to TAR RNA in the U-rich RNA bulge region and this bindig facilitates the interactions of the CycT1-Tat heterodimer at the other two sites in the RNA loop region. On the basis of our results, we suggest a model where CycT1 interacts with Tat protein and positions the protein complex to make contacts with the G34 region of the loop sequence; G34 is critical for CycT1-Tat binding and forms a C30 ident dot G34 base pair. Two functional groups, O6 and N7, at nucleotide positions 32 and 34 in the TAR loop are essential for CycT1-Tat intrractions with TAR RNA. The identity of two nucleotides, U31 and G33, is not critical, but they contribute to the stabilization of the RNA-protein complex. The presence of a single-nucleotide bulge of A35 or C35 is essential for distortion of the backbone RNA structure as well as the accessibility of functional groups in the major groove of the double-helical region. CycT1-Tat interaction with TAR RNA represents another example of the flexibility and complexity of RNA structure involved in protein recognition.
机译：人类免疫缺陷病毒的复制需要Tat蛋白，该蛋白通过与阳性转录延伸因子复合物（P-TEFvb）的细胞周期蛋白T1（CycT1）亚基相互作用，在HIV-1启动子上激活RNA聚合酶II转录的延长。 Tat通过其反式激活域直接与P-TEFbv的CycTR1 suvbunit结合，并诱导P-TEFb环序列特异性结合到新生的HIV-1 TAR RNA上。通过使用凝胶电泳方法和一整套TAR环突变体，我们已经确定了高亲和力CycT1-Tat-TAR三元复合物形成的序列和结构决定因素。我们的结果表明CycT1和Tat与TAR RNA的结合是高度合作的，观察到其容量为85％，希尔系数为2.7，解离常数（K_D）为2.45 nM。这些结果表明在TAR RNA上有三个结合位点。在没有Tat的情况下，CycT1不会结合TAR RNA，尽管Tat与TAR的结合是可检测的，但在没有CycT1的情况下却效率很低。可以想象，CycT1-Tat异二聚体直接与富含U的RNA凸起区域中的TAR RNA结合，并且这种结合促进了CycT1-Tat异二聚体在RNA环区域中其他两个位点的相互作用。根据我们的结果，我们提出了一个CycT1与Tat蛋白质相互作用并将该蛋白质复合物定位成与环序列的G34区接触的模型。 G34对于CycT1-Tat结合至关重要，并形成一个C30身份点G34碱基对。 TAR环中第32和34位核苷酸的两个官能团O6和N7对于用TAR RNA进行CycT1-Tat插入至关重要。 U31和G33这两个核苷酸的身份并不重要，但它们有助于稳定RNA-蛋白质复合物。 A35或C35的单核苷酸凸起对于骨架RNA结构的扭曲以及双螺旋区主沟中官能团的可及性至关重要。 CycT1-Tat与TAR RNA的相互作用代表了参与蛋白质识别的RNA结构的灵活性和复杂性的另一个例子。

著录项

相似文献

  • 外文文献
  • 专利
获取原文

客服邮箱：kefu@zhangqiaokeyan.com

京公网安备：11010802029741号 ICP备案号：京ICP备15016152号-6 六维联合信息科技 (北京) 有限公司©版权所有

  • 客服微信

  • 服务号