COBRE: NDSU: PROJECT 5: NEW INVESTIGATOR

这个子项目是许多利用资源的研究子项目之一 由NIH/NCRR资助的中心拨款提供。子项目的主要支持 而子项目的主要调查员可能是由其他来源提供的， 包括其它NIH来源。 列出的子项目总成本可能 代表子项目使用的中心基础设施的估计数量， 而不是由NCRR赠款提供给子项目或子项目工作人员的直接资金。 A节：具体目标 我实验室研究的最终目标是利用各种结构和生物化学 技术来理解蛋白质功能的原子基础， （降解细胞内物质的溶酶体靶向途径）和先天免疫 途径;这些途径之间的串扰;这些途径/蛋白质在 对病原体的防御;以及不同病原体对这些蛋白质的抑制。 一 项目重点研究Beclin 1的结构、功能和分子机制， 一种重要的自噬效应物 我们之前的工作，部分由NIH R21基金资助， 研究了细胞和病毒Bcl-2蛋白抑制Beclin的分子机制 1、自噬 我们计划继续研究Beclin 1蛋白，包括其 与其他自噬蛋白和病原体编码蛋白的相互作用。 另一 项目的重点是Rig-I，抗病毒先天免疫的关键介质。 我们计划调查 Rig-I在先天防御中作用的原子基础，通过研究分子内， Rig-I CARD结构域的通路或跨通路相互作用， 触发或抑制各种途径。 所有这些研究的先决条件是获得纯的、均质的、重组的 蛋白 我们正在使用COBRE资金生产这些试剂并获得初步数据 申请其他赠款，以进一步研究这些途径。 COBRE的具体目标 资金用于大量过量表达和纯化下列重组蛋白 适用于随后的结构和生物化学研究： + 目的1：不同的Beclin 1结构域。 + 目标2：Rig-I CARD结构域。 + 目的3：IPS-1的CARD结构域，其下游信号传导伙伴。 + 目的4：Rig-I和LGP 2（Rig-I同源物）的阻遏结构域。 + 目的5：与Rig-I和IPS-1相互作用的重要自噬效应子Atg 5 CARD域。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Section A: Specific Aims The ultimate goal of research in my laboratory is to use various structural and biochemical techniques to understand the atomic basis of function of proteins of pathways like autophagy (a lysosomal targeting pathway for degradation of intracellular objects) and innate immune pathway; the cross-talk between these pathways; the role of these pathways/proteins in the defense against pathogens; and the inhibition of these proteins by different pathogens. One project focuses on investigating the structure, function and molecular mechanism of Beclin1, an essential autophagy effector. Our previous work, funded in part by an NIH R21 grant, investigated the molecular mechanism by which cellular and viral Bcl-2 proteins inhibit Beclin 1 and autophagy. We plan to continue our investigation of the Beclin 1 protein, including its interaction with other autophagy proteins and with pathogen-encoded proteins. Another project focuses on Rig-I, a key mediator of anti-viral innate immunity. We plan to investigate the atomic basis of the role of Rig-I in innate defenses by studying the intra-molecular, intra- pathway or cross-pathway interactions of the Rig-I CARD domains that transmit signals to trigger or repress various pathways. A prerequisite of all these studies is the availability of pure, homogeneous, recombinant protein. We are using COBRE funding to produce these reagents and obtain preliminary data to apply for other grants to further investigate these pathways. Specific aims of the COBRE funding are to overexpress and purify the recombinant proteins listed below, in quantities suitable for subsequent structural and biochemical studies: + Aim 1: Different Beclin 1 domains. + Aim 2: Rig-I CARD domains. + Aim 3: CARD domains of IPS-1, its downstream signaling partner. + Aim 4: Repressor domains of Rig-I and LGP2 (a Rig-I homolog). + Aim 5: The essential autophagy effector Atg5, which interacts with the Rig-I and IPS-1 CARD domains.