X-RAY DIFFRACTION ANALYSIS OF METABOLITE SENSING RIBOSWITCHES, G-PROTEIN/SMALL M

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 这项提议的目标是收集与生物学和公共健康相关的三种不同生物大分子的高分辨率X射线衍射数据。我们的第一个目标是学习细菌前Q1核糖开关识别代谢物的模式。这些信息对于理解这一新的基因调控范例至关重要，因为它可能成为抗菌靶标。以前，我们在SSRL确定了PreQ1核糖开关的结构，现在已经获得了在各种代谢物存在下生长的新晶体，这些代谢物跨越了广泛的结合亲和力范围(Kd=5 nM到500 nM)。我们还在研究G蛋白β-伽马与各种小分子的结合模式，这些小分子被我们的合作者证明是影响细胞信号传递的。这些分子被假定结合在贝塔亚基上的一个“热点”上。我们的目标是通过差分傅立叶技术定位这些分子，这对开发新的G蛋白信号效应器具有重要意义。最后，我们正在努力确定HIV-1蛋白Vif与库林环连合酶复合体的晶体结构。这种病毒/宿主蛋白的相互作用还没有在分子水平上得到表征。我们的目标是鉴定衍射性良好的晶体，以便进行结构测定。我们的复合体包括两种人类蛋白，EloB和EloC，以及HIV-1 Vif。

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. The goal of this proposal is to collect high-resolution X-ray diffraction data for three different biomacromolecules that are relevant to biology and public health. Our first objective will be to learn the mode of metabolite recognition by a bacterial, PreQ1 riboswitch. Such information is essential to understand this novel gene regulatory paradigm, which may serve as an antimicrobial target. Previously we determined the structure of the PreQ1 riboswitch at SSRL and have now obtained new crystals grown in the presence of various metabolites that span a broad range of binding affinities (Kd = 5 nM to 500 nM). We are also investigating the mode of binding of the G-protein beta-gamma to various small molecules shown to affect cell signaling by our collaborator. These molecules are hypothesized to bind in a "hotspot" on the beta subunit. Our goal is to locate this molecules by difference Fourier techniques, which has implications for developing novel effectors of G-protein signaling. Finally, we are working to determine the crystal structure of the HIV-1 protein Vif in complex with components of the Cullin-RING ligase complex. This viral/host protein interaction has not been characterized at the molecular level. Our goal is to identify well-diffracting crystals in order to proceed with a structure determination. Our complex comprises two human proteins, EloB and EloC, as well as HIV-1 Vif.