IRON SUBSTITUTED ZINC FINGER PROTEINS

蛋白质折叠和错误折叠是一个根本性的问题 生物化学。自然态是怎样的(推定的热力学 基态)在生理时间尺度上达到的；尤其是如何 错误折叠状态(“运动陷阱”)被避免了吗？什么是 蛋白质折叠中间体的结构？有没有 “蛋白质折叠密码”，如果是这样的话，独特的“折叠性”是普遍的吗？ 蛋白质序列的性质？这些问题长期以来一直是 对物理、生物化学和生物物理学感兴趣。的最新进展 分子医学现在强调了它们与 人类疾病的发病机制。我们打算解剖一位祖先 蛋白质折叠途径：胰岛素-IGF基序的折叠途径。初步 研究证实，胰岛素样生长的氧化折叠 因子1(IGF-1)在以下条件下产生两种产物(二硫键异构体) 热力学控制。将进行多维核磁共振研究 以确定其与天然IGF-1异构体的结构关系 胰岛素样生长因子-1与胰岛素的一种新的错误折叠状态(自然结构 生物学2,129-138)。基因工程类似物和多肽 蛋白质折叠中间体的模型将被设计来测试 以下命题：假设1.错误折叠的核磁共振研究 状态(动力学陷阱)将确定天然蛋白质的决定因素 折叠；假设2。同源折叠的比较研究 途径(胰岛素原和IGF-1)将识别保守的中间体 和过渡态；以及假设3。非随机路径 二硫键的形成将使多肽和 蛋白质折叠中间体的重组模型。特别的 感兴趣的将是合作使用非标准氨基酸 固相法合成多肽构建新的多肽模型 蛋白质折叠中间体。通过结合2D和多维 核磁共振波谱与现代化学合成和遗传学方法 工程学，这一应用程序提供了令人兴奋的可能性 描绘了从蛋白质序列到结构的路径。

Protein folding and misfolding represent a fundamental problem in biochemistry. How is the native state (the presumed thermodynamic ground state) reached on a physiologic timescale; in particular, how are misfolded states ("kinetic traps") avoided? What are the structures of protein-folding intermediates? Is there a "protein-folding code", and if so, is unique "foldability" a general property of protein sequences? These questions have long been of interest in physical biochemistry and biophysics. Recent advances in molecular medicine now highlight their central relevance to the pathogenesis of human diseases. We propose to dissect an ancestral protein-folding pathway: that of the insulin-IGF motif. Preliminary studies have established that oxidative folding of insulin-like growth factor 1 (IGF-1) yields two products (disulfide isomers) under thermodynamic control. Multidimensional NMR studies will be conducted of the IGF-1 isomer to determine its structural relationship to native IGF-1 and to a novel misfolded state of insulin (Nature Structural Biology 2, 129-138). Genetically engineered analogs and peptide models of protein-folding intermediates will be designed to test the following propositions:Hypothesis 1. That NMR studies of misfolded states (kinetic traps) will identify determinants of native protein folding; Hypothesis 2. That comparative studies of homologous folding pathways (proinsulin and IGF-1) will identify conserved intermediates and transition states; and Hypothesis 3. That the non-random pathway of disulfide bond formation will enable design of peptide and recombinant models of protein-folding intermediates. Of particular interest will be collaborative use of non-standard amino acids in solid-phase peptide synthesis to construct novel peptide models of protein-folding intermediates. By combining 2D and multidimensional NMR spectroscopy with modern methods of chemical synthesis and genetic engineering, this application offers the exciting possibility of delineating a pathway from protein sequence to structure.