IRON SUBSTITUTED ZINC FINGER PROTEINS

蛋白质折叠和错误折叠代表了一个基本问题， 生物化学 自然状态（假定的热力学 基态）在生理时间尺度上达到;特别是，如何 是否避免了错误折叠状态（“动力学陷阱”）？ 有哪些 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，是唯一的“折叠性”的一般 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感兴趣。 的最新进展 分子医学现在强调了它们与人类的核心相关性， 人类疾病的发病机制。 我们打算解剖一个祖先 蛋白质折叠途径：胰岛素-IGF基序。 初步 研究已经确定，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（二硫化物异构体）： 热力学控制 将进行多维核磁共振研究 的IGF-1异构体，以确定其结构关系， IGF-1和胰岛素的新错误折叠状态（Nature Structural Biology 2，129-138）。 基因工程类似物和肽 将设计蛋白质折叠中间体模型来测试 假设：假设1。 核磁共振研究错误折叠的 状态（动力学陷阱）将识别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）将确定保守的中间体 和过渡状态;和假设3。 非随机路径 二硫键的形成将使肽的设计， 蛋白质折叠中间体的重组模型。 特别 感兴趣将是非标准氨基酸的协作使用， 固相肽合成，以构建新的肽模型， 蛋白质折叠中间体。 通过结合二维和多维 核磁共振光谱与现代化学合成和遗传方法 工程，该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.