STRUCTURAL STUDIES OF RED CELL FERRITINS

红细胞铁蛋白的结构研究

• 批准号: 2414917
• 负责人: NORMA M. ALLEWELL
• 金额: $ 9.83万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-05-01 至 2000-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2414917
• 关键词: Mossbauer spectrometry; Raman spectrometry; X ray crystallography; atomic absorption spectrometry; chemical binding; chemical models; computer simulation; erythrocytes; ferritin; molecular site; mutant; protein sequence; protein structure function; recombinant proteins

Ferritin is a multisubunit protein that concentrates and sequesters as many as 4,500 iron atoms within a hollow protein shell in animal, plant and bacterial cells. There are at least three ferritin genes in animals that differ in expression in different tissues, under different environment conditions and with development; their rates and mechanisms of iron accumulation, storage and turnover are also different. Our goal is to investigate the mechanism of action of ferritin and the structural basis of these functional differences by using X-ray crystallography to determine and compare the three-dimensional structures of recombinant amphibian red H and L ferritins and a set of single site mutants with altered function under different experimental conditions. Particular emphasis will be placed on mutants that eliminate or restore the ability to form the early iron-tyrosine intermediate that forms only in H- ferritin, mutations involving a set of mobile sidechains associated with a spine of hydration that we have identified in previous studies and mutations near the symmetry axes. We will also use a package developed in this laboratory specifically for modeling large macromolecular assemblies to model and compare electrostatic effects in the various structures. The functional properties of the various proteins will be determined and compared in solution, principally by EXAFS, resonance Raman, Mossbauer and uv-visible absorption spectroscopy. The basis of this proposal is our earlier studies of wild type amphibian red cell L-ferritin and two mutants which indicated that the ferritin structure is more plastic than had previously been appreciated and can be differentially solvated. We wish to elucidate the rules that govern these structural changes and their relationship to function, particularly the differences in L- and H- ferritins. Since the subunits of ferritin are four-helix bundles, this study will provide insight into how this common structural motif in proteins accommodates to changes in sequence and external conditions. The mechanism of action of ferritin is relevant to understanding the molecular basis of iron metabolism in normal health and in diseases such as thalassemia, hemochromatosis and sickle cell anemia or other conditions requiring hypertransfusion or hemodialysis.

铁蛋白是一种多亚基蛋白，将其集中和隔离为 动物，植物中的空心蛋白壳中的多达4,500个铁原子 和细菌细胞。 动物中至少有三个铁蛋白基因 在不同组织中的表达不同，在不同的组织下 环境状况和发展；他们的费率和机制 铁的积累，存储和营业额也不同。 我们的目标是 研究铁蛋白和结构的作用机理 通过使用X射线晶体学到 确定并比较重组的三维结构 两栖红H和L铁蛋白以及一组单位突变体 在不同的实验条件下功能改变。 特别的 重点将放在消除或恢复能力的突变体上 形成仅在H-中形成的早期铁酪氨酸中间体 铁蛋白，涉及与一套移动侧技术相关的突变 我们在以前的研究中已经确定的水合脊柱 对称轴附近的突变。 我们还将使用开发的软件包 该实验室专门为大型大分子组件建模 建模和比较各种结构中的静电效应。 这 将确定各种蛋白质的功能特性，并 在解决方案中，主要由Exafs，Resonance Raman，Mossbauer和 紫外可见的吸收光谱。 该提议的基础是我们的 野生型两栖红细胞L-铁素和两个突变体的早期研究 这表明铁蛋白结构的塑性比 以前已被欣赏，可以差异化。 我们希望 阐明管理这些结构性变化及其的规则 与功能的关系，尤其是L-和H-的差异 铁蛋白。 由于铁蛋白的亚基是四螺旋束，所以 研究将提供有关如何在这种常见结构主题中的见解 蛋白质适应序列和外部条件的变化。 这 铁蛋白的作用机理与理解分子有关 铁代谢在正常健康和疾病中的基础 地中海贫血，血色素瘤病和镰状细胞贫血或其他疾病 需要大量灌注或血液透析。