STRUCTURE & FUNCTION OF THE SV40 T-AG DNA BINDING DOMAIN

DNA复制的启动是一个复杂的过程， 在非常广泛的意义上。目前，最好的真核生物模型 DNA复制起始的研究是猿猴病毒40（SV4O）， 体外复制系统启动SV4O复制需要单个 称为T抗原（T-ag）的病毒蛋白。www.example.com发挥的关键作用 包括与SV4O的位点特异性结合 起源、随后解旋事件的催化和 启动DNA合成所必需的额外蛋白质。然而，在这方面， 我们对这些事件的了解是有限的，因为缺乏结构性的 关于T-ag或T-ag DNA结合结构域（T-ag-bd）的信息。的 为解决这些问题，提出了以下具体目标。 I.确定SV40 T抗原DNA结合域的结构。 二.为了表征T-ag-bd的生化特性， 某些突变形式的T-ag-bd，并检查结构的 那些具有有趣特性的T-ag-bd突变体。 三.描述T-ag-bd与DNA的相互作用。 本申请中提出的工作从一个基础科学的角度来说是有意义的 因为它将提供蛋白质结构域的第一个结构 识别复制起点，无论是来自原核生物还是 真核生物来源。此外，现有序列数据表明T-ag-bd 与其他已知的DNA结合蛋白无关，这表明 结构可能揭示了一个新的蛋白质结构基序。对此的了解 结构，或与DNA复合的结构域的结构，将 为我们进一步理解复制做出了重大贡献， 揭示蛋白质/DNA接触，使特异性识别， 与复制起点结合。 所提议的工作的健康相关意义源于以下事实： SV40 T-ag与BK和JC编码的T-ag非常同源 病毒。这些病毒在人类中诱发多种疾病，包括 癌例如，JC病毒诱导进行性多灶性 脑白质病，一种存在于许多艾滋病患者中的疾病。

Initiation of DNA replication is a complicated process that is understood in only very broad terms. At present, the best eukaryotic model for studies of initiation of DNA replication is the Simian Virus 40 (SV4O) in vitro replication system. Initiation of SV4O replication requires a single viral protein termed T-antigen (T-ag). Critical roles played by T.ag during initiation of replication include site specific binding to the SV4O origin, catalysis of subsequent unwinding events and recruitment of additional proteins necessary for initiation of DNA synthesis. However, our understanding of these events is limited owing to a lack of structural information about T-ag or the T-ag DNA binding domain (T-ag-bd). The following specific aims are proposed to address these issues. I. To determine the structure of the DNA binding domain of SV4O T-antigen. II. To characterize the biochemical properties of the T-ag-bd, and of certain mutant forms of the T-ag-bd, and to examine the structures of those mutant forms of the T-ag-bd with interesting properties. III. To delineate the interaction of the T-ag-bd with DNA. The work proposed in this application is significant from a basic science standpoint because it will provide the first structure of a protein domain that recognizes an origin of replication, whether from a prokaryotic or eukaryotic source. Moreover, existing sequence data indicate the T-ag-bd is not related to other known DNA binding proteins, suggesting the structure may reveal a new protein structural motif. A knowledge of this structure, or of the structure of this domain complexed to DNA, will contribute significantly to advancing our understanding of replication by revealing the protein/DNA contacts that enable specific recognition and binding to an origin of replication. The health related significance of the work proposed stems from the fact that SV4O T-ag is very homologous to the T-ags encoded by the BK and JC viruses. These viruses induce a number of diseases in humans, including cancer. For example, JC virus induces progressive multifocal leukoencephalopathy, a disease present in many AIDS patients.