MIMICS OF HELIX TURN HELIX PEPTIDES

螺旋转角螺旋肽的模拟物

• 批准号: 6180628
• 负责人: FELICIA A ETZKORN
• 金额: $ 4.37万
• 依托单位: UNIVERSITY OF VIRGINIA
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-05-01 至 2000-08-09
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6180628
• 关键词: DNA binding protein; chimeric proteins; conformation; crosslink; nuclear magnetic resonance spectroscopy; peptide analog; peptide chemical synthesis; protein engineering; protein folding; protein structure; protein structure function; stereochemistry; structural biology; transcription factor

DESCRIPTION: The goal of this project is to mimic the helix-turn-helix (HTH) motif of the POU Oct-1 homeodomain and the 434 Cro repressor by constraining the native conformation of this small portion of the protein. The helix-turn-helix motif is found in several DNA-binding proteins that have known 3-dimensional structures. Such chimeric molecules can serve a two-fold health-related purpose. First, the design and biochemical evaluation of the chimeric molecules will explore fundamental aspects of protein folding and protein-DNA interactions. Second, each of the mimics described could serve as biochemical probes of developmental processes. POU homeodomains are transcription factors for human growth hormone, histones, snRNAs, immunoglobulin, herpes simplex virus and other medically important genes. The PI proposes to constrain the conformation of peptides corresponding to the HTH motif via covalent bonds between buried hydrophobic side-chains. Two approaches to this unusual type of side-chain linkage will be taken: 1) aryl linkage and 2) alkyl linkage. The templates are designed to stabilize the conformation of the flexible turn. The designs feature stable organic side-chains which will favor protein folding by burial of the additional hydrophobic moiety. Each template will be synthesized stereospecifically from amino acid starting materials and incorporated by solid-phase synthesis into a peptide corresponding to the sequence of the structurally well-characterized Cro repressor and POU Oct-1 homeodomain. Peptide engineering of the alpha-helices will introduce salt bridges and residues with high helix propensities to improve the helicity of attached peptides. A helix-to-helix crosslink will be introduced to improve the tertiary structure stability. The HTH mimics will be characterized chemically (HPLC, MS, NMR), structurally (CD, NMR and/or x-ray crystallography) and functionally (DNA-binding affinity) to ascertain the success of the mimics.

描述：该项目的目的是模仿螺旋 - 螺旋 - 螺旋 （HTH）POU OCT-1同源域和434 CRO抑制器的图案 限制蛋白质这一小部分的天然构象。 在几种DNA结合蛋白中发现螺旋 - 转移螺旋序列 已知三维结构。 这样的嵌合分子可以使用 两倍与健康相关的目的。 首先，设计和生化 嵌合分子的评估将探讨 蛋白质折叠和蛋白-DNA相互作用。 第二，每个模仿 所描述的可以用作发育过程的生化探针。 pou 同源域是人类生长激素，组蛋白的转录因子， SNRNA，免疫球蛋白，单纯疱疹病毒和其他重要的医学重要 基因。 PI提议约束肽的构象 通过埋葬疏水之间的共价键对应于HTH基序 侧链。 这种不寻常类型的侧链连接的两种方法将 采取：1）芳基连接和2）烷基连接。 模板是设计的 稳定柔性转弯的构象。 设计功能 稳定的有机侧链将有利于埋葬的蛋白质折叠 额外的疏水部分。 每个模板将合成 立体特定于氨基酸起始材料，并通过 固相合成到对应于与序列相对应的肽中 结构良好的CRO抑制剂和POU OCT-1同源域。 Alpha螺旋的肽工程将引入盐桥和 具有高螺旋倾向的残留物以改善附着的螺旋性 肽。 将引入一个螺旋到螺旋交叉链接以改善 三级结构稳定性。 HTH模仿将被描述 化学（HPLC，MS，NMR），结构上（CD，NMR和/或X射线 晶体学）和功能（DNA结合亲和力）以确定 模仿的成功。