BIOCHEMISTRY OF HEPATITIS C VIRUS REPLICASE

描述（由申请人提供） 拟议研究的长期目标是了解和定义 HCV复制酶的生物化学和结构方面。这将需要 鉴定氨基酸残基，以及特定的结构域上的 参与病毒RNA基因组复制过程的酶分子。 最近解决的丙型肝炎病毒复制酶的三维晶体结构， 结合分子模拟，已经确定了一些氨基酸 疑似参与酶促反应特定步骤的残留物。 这些残基将被靶向用于定点诱变，并且所得到的突变体将被靶向。 突变酶的特征在于它们的生物化学性质 性质，以阐明其在催化反应中的功能作用。 过程沿着假定的RNA结合轨道进行沿着的诱变研究将阐明 这些残基在RNA复制过程中的作用。这些研究 将直接应用于定义一些分子的侧链取向 催化结构域中的氨基酸残基。知识大全 HCV复制酶的功能性三维结构将有助于更好地理解 RNA依赖性RNA合成的机制及抗 HCV复制酶药物。这些信息的提供反过来将进一步 促进开发新的HCV-复制酶导向的特异性原子 这些尺寸将作用于HCV复制酶的特定结构元件。的 提出了以下具体目标。 目的1：丙型肝炎病毒复制酶的生物化学特性。目的2：阐明 特定氨基酸残基在聚合酶功能中的催化作用。目的 3：确定HCV复制酶与其他蛋白质之间的蛋白质-蛋白质相互作用 非结构性病毒蛋白。

DESCRIPTION (provided by applicant) The long-term goal of the proposed research is to understand and define the biochemical and structural aspects of HCV replicase. This would necessitate identification of the amino acid residues, as well as specific domains on the enzyme molecule involved in the process of copying the viral RNA genome. Recently solved three-dimensional crystal structures of HCV replicase in conjunction with molecular modeling have pinpointed a number of amino acid residues suspected to be involved in specific steps of the enzymatic reaction. These residues will be targeted for site directed mutagenesis and the resulting mutant enzymes will be characterized with respect to their biochemical properties in order to elucidate their functional roles in the catalytic process. Mutagenesis studies along the putative RNA binding track will clarify the roles of these residues in the process of RNA replication. These studies will have direct application in defining the side chain orientation of a number of amino acid residues in the catalytic domain. Knowledge of the complete functional 3-D structure of HCV replicase will help in better understanding of the mechanisms of RNA dependent RNA synthesis and inhibitory action of anti HCV-replicase drugs. Availability of this information, in turn, will further facilitate development of new HCV-replicase directed agents of specific atomic dimensions that would act at specific structural elements of HCV-replicase. The following specific aims are proposed. Aim 1: Biochemical characterization of HCV replicase. Aim 2: To elucidate the catalytic roles of specific amino acid residues in the polymerase function. Aim 3: To determine protein-protein interactions between HCV replicase and other nonstructural viral proteins.