STRUCTURAL STUDIES ON GENE V PROTEIN

这个项目的目标是了解蛋白质是如何结合到 了解和开发单链核酸 经常由多种氨基酸变化引起的效应的可加性 蛋白质的结构和功能。单链核酸- 结合蛋白在关键的细胞过程中发挥作用，如DNA RNA翻译的复制、重组和控制。如果这些方法 其中蛋白质与单链核酸相互作用 详细了解，则有可能修改这些函数 以一种靶向方式对这些类别的蛋白质进行识别。这将是 在治疗由于身体功能缺陷引起的疾病方面很重要 这些蛋白质的功能。蛋白质是非常复杂的大分子， 以一种完全理性的方式设计它们的属性 异常困难，因为改变氨基酸的影响 蛋白质的序列不能被详细预测。如果功能正常且 蛋白质中氨基酸取代的结构效应可能是 对某些类别的多重突变的准确预测 组成单一氨基酸替换的影响，然后这个 流程可以简化。这样的简化可能会大大减少 获得所需蛋白质所需的时间和实验 一组特征，例如对 特定的核酸序列。这个项目的第一部分是 旨在了解基因V蛋白是如何与单个- 以及该蛋白质是如何优先识别 RNA和同源DNA的特定序列。这是要完成的 通过测定基因V之间形成的络合物的晶体结构 特异性和非特异性结合的蛋白质和寡核苷酸 到蛋白质。我们已经获得了两种高质量的共晶 基因V蛋白-寡核苷酸复合体将X射线衍射到 分辨率分别为3.0埃和3.4埃。目标是 项目的第二部分是为了扩大我们对 突变对化合物结构和性质影响的可加性 蛋白。对可加性的这种理解是通过研究 基因V蛋白单突变和双突变对大鼠血管紧张素转换酶活性的影响 蛋白质的结构和性质，重点研究了它们之间的关系 在两个突变效应的可加性程度和 当单独发生突变时，受突变影响的区域。 该项目的第三部分旨在开发以下技术 将使大分子结晶学实验得以分析 更准确地说。我们预计这个项目的结果将会有一个 在生物技术领域和在治疗中的重大影响 人类疾病。

The goals of this project are to understand how a protein binds to single-stranded nucleic acids and to understand and exploit the additivity of effects often resulting from multiple amino acid changes in a protein on its structure and function. Single-stranded nucleic acid- binding proteins play roles in key cellular processes such as DNA replication, recombination, and control of RNA translation. If the ways in which proteins interact with single-stranded nucleic acids were understood in detail, then it might be possible to modify the functions of these classes of proteins in a target fashion. This would be important in the treatment of diseases due to deficiencies in the function of these proteins. Proteins are very complex macromolecules, and engineering their properties in a completely rational manner is exceptionally difficult because the effects of changing the amino acid sequence of a protein cannot be predicted in detail. If functional and structural effects of amino acid substitutions in a protein could be accurately predicted for certain classes of multiple mutations based on the effects of the constituent single amino acid substitutions, then this process could be simplified. Such a simplification could greatly reduce the time and experimentation required to obtain a protein with a desired set of characteristics, such as a certain stability and affinity for a particular sequence of nucleic acid. The first part of this project is directed towards understanding how gene V protein interacts with single- stranded nucleic acids and how the protein preferentially recognizes a specific sequence of RNA and the cognate DNA. This is to be accomplished by determining crystal structures of complexes formed between gene V protein and oligonucleotides that bind specifically and non-specifically to the protein. We have already obtained high-quality co-crystals of two gene V protein-oligonucleotide complexes that diffract X-rays to resolutions of 3.0 angstroms and 3.4 angstroms, respectively. The goal of the second part of the project is to broaden our understanding of the additivity of effects of mutations on the structure and properties of a protein. This understanding of additivity is to be obtained by examining the effects of single and double mutations in gene V protein on the structure and properties of the protein, focusing on the relationship between the extent of additivity of effects of two mutations and the regions structurally affected by the mutations when made individually. The third part of this project is aimed at developing techniques that will allow macromolecular crystallographic experiments to be analyzed more accurately. We expect that the results of this project will have a substantial impact in the field of biotechnology and in the treatment of human disease.