Beta Sheet Formation in a Simple Model System

描述（由申请人提供）：我们正在研究稳定蛋白质的相互作用的能量学和物理化学基础。这些研究包括分析溶液中小单体13层蛋白的稳定性，以及侧链-侧链相互作用在体内和体外调节蛋白质折叠速率中的作用。此外，我们的研究还探讨了决定蛋白质在溶液中是否可溶或形成淀粉样原纤维的能量平衡和折叠机制。我们还在开发x射线衍射方法，以获得淀粉样蛋白原纤维的高分辨率结构信息。我们在这些研究中使用的蛋白质是igg结合蛋白G （beta1）和绿色荧光蛋白（GFP） B1结构域的一系列变体。我们对绿色荧光蛋白的研究使我们能够设计出一种新的系统，它对检测蛋白质之间的相互作用具有很大的效用。我们把绿色荧光蛋白分成两半，只有融合到两个相互作用的蛋白质上才能重新组装。我们正在使用这个系统来研究蛋白质-蛋白质相互作用的亲和力和特异性。提出了四肽重复（TPR）介导的蛋白-蛋白相互作用的具体例子。分裂GFP系统还提供了一种方法，通过它来识别“孤儿蛋白”的结合伙伴，其相互作用伙伴尚不清楚。

DESCRIPTION (provided by applicant): We are investigating the energetics and physical-chemical basis of the interactions that stabilize proteins. These studies include analyses of the stability of small, monomeric 13-sheet proteins in solution and also the role that sidechain-sidechain interactions play in modulating the rate of protein folding both in vivo and in vitro. In addition, our studies investigate the energetic balance and folding mechanism that determines whether a protein will be soluble or form amyloid fibrils in solution. We are also developing x-ray diffraction methods to gain high-resolution structural information on amyloid fibrils. The proteins we use for these studies are a series of variants of the B1 domain of IgG-binding protein G (beta1) and the Green Fluorescent Protein (GFP). Our studies with GFP have allowed us to design a novel system that has great utility for the detection of protein-protein interactions. We have dissected GFP into two halves that can only be reassembled if fused to two interacting proteins. We are using this system to investigate the affinity and specificity of protein-protein interactions. The specific example of tetratrcopeptide repeat (TPR) mediated protein-protein interactions is presented. The split GFP system also provides a means by which to identify binding partners for "orphan proteins" whose interaction partners are not yet known.