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STRUCTURAL STUDIES ON GENE V PROTEIN

V 基因蛋白质的结构研究

基本信息

批准号：
6018702
负责人：
THOMAS C. TERWILLIGER
金额：
$ 26.82万
依托单位：
UNIVERSITY OF CALIF-LOS ALAMOS NAT LAB
依托单位国家：
美国
项目类别：
财政年份：
1987
资助国家：
美国
起止时间：
1987-07-01 至 2000-12-31
项目状态：
已结题

项目摘要

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.

该项目的目标是了解蛋白质如何结合到单链核酸，并了解和利用作用的加和性，通常由多个氨基酸的变化引起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，复制、重组和RNA翻译的控制。如果方法其中与单链核酸相互作用的蛋白质被详细了解，那么就有可能修改功能这些蛋白质的靶点。这将是重要的是在治疗疾病，由于缺乏在这些蛋白质的功能。蛋白质是非常复杂的大分子，并以完全合理的方式设计它们的属性，因为改变氨基酸的影响，蛋白质的序列无法详细预测。如果功能正常，蛋白质中氨基酸取代的结构效应可能是准确预测某些类别的多个突变，组成的单一氨基酸取代的影响，那么这个过程可以简化。这种简化可以大大减少获得具有所需蛋白质的时间和实验一组特征，例如对一个人的某种稳定性和亲和力，特定的核酸序列。这个项目的第一部分是旨在了解基因V蛋白如何与单个链核酸以及蛋白质如何优先识别 RNA和同源DNA的特定序列。这是要完成的通过测定基因V和基因C之间形成的复合物的晶体结构，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蛋白质-寡核苷酸复合物，分辨率分别为3.0埃和3.4埃。目标项目的第二部分是扩大我们对突变对结构和性质影响的加和性蛋白这种对可加性的理解是通过考察基因V蛋白的单突变和双突变对蛋白质的结构和性质，重点是关系两个突变的效果的加和性程度与当单独进行时，结构上受突变影响的区域。该项目的第三部分旨在开发技术，将允许分析大分子晶体学实验更准确地说。我们预计，该项目的结果将有一个在生物技术领域和在治疗人类疾病。