NS3 HELICASE

该子项目是利用该技术的众多研究子项目之一 资源由 NIH/NCRR 资助的中心拨款提供。子项目及 研究者 (PI) 可能已从 NIH 的另一个来源获得主要资金， 因此可以在其他 CRISP 条目中表示。列出的机构是 对于中心来说，它不一定是研究者的机构。 RNA 代谢需要大量 ATP 依赖性分子马达蛋白的作用，这些蛋白被认为可以运输、重塑和解开 RNA 序列中的二级结构。 许多这些分子马达是死盒或密切相关的蛋白质。 许多正链 RNA 病毒，例如丙型肝炎病毒 (HCV)，需要这些蛋白质的活性来进行 RNA 复制。 确定这些蛋白质的机制及其具体功能对于我们理解病毒复制和 RNA 代谢具有重要意义。非结构蛋白 3 (NS3) 是一种 RNA 马达蛋白（或解旋酶），是 HCV 复制所必需的。 我们的数据支持这样的假设：NS3 在单体和寡聚体之间处于平衡状态，并且其 RNA 解旋活性随着寡聚化的增加而增加。 我们的数据表明 NS3 解旋酶可以在 RNA 和 DNA 上形成丝。 我们希望建立合作伙伴关系来可视化这些细丝。 随附的手稿描述了我们关于 DNA 丝形成的一些证据。 然而，大多数支持细丝形成的数据尚未公布。 我们希望建立这种合作，以便彻底研究 NS3 的蛋白质-蛋白质和蛋白质-DNA 相互作用。我们没有 EM 方面的经验，但知道许多用这种方法表征细丝形成的例子。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. RNA metabolism requires the action of numerous ATP-dependent, molecular motor proteins that are believed to transport, remodel, and unwind secondary structures in RNA sequences. Many of these molecular motors are DEAD-box or closely related proteins. Many positive strand RNA viruses, such as the hepatitis C virus (HCV), require the activity of these proteins for RNA replication. Determination of the mechanism of these proteins and their specific functions is of fundamental importance to our understanding of viral replication as well as RNA metabolism in general. Non-structural protein 3 (NS3) is an RNA motor protein (or helicase), that is necessary for HCV replication. Our data supports the hypothesis that NS3 exists in equilibrium between monomeric and oligomeric species, and that its RNA unwinding activity increases with increasing oligomerization. Our data suggest that NS3 helicase can form filaments on RNA and DNA. We wish to establish a collaborative effort to visualize these filaments. The attached manuscripts describes some of our evidence for filament formation on DNA. However, most of the data supporting filament formation is yet to be published. We wish to establish this collaboration in order to thoroughly investigate protein-protien and protein-DNA interactions of NS3. We have no experience in EM, but are aware of numerous examples in which filament formation has been characterized by this method.