STRUCTURES INFLUENCE ON REACTIVITY IN METALLOENZYMES

结构对金属酶反应性的影响

• 批准号: 6386177
• 负责人: Julia A Kovacs
• 金额: $ 23.77万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-02-01 至 2002-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6386177
• 关键词: X ray crystallography; active sites; alcohol dehydrogenase; cobalt; cyanides; cysteine; electron spin resonance spectroscopy; enzyme activity; enzyme complex; enzyme inhibitors; enzyme mechanism; enzyme model; enzyme structure; enzyme substrate; hydro lyase; hydrogenase; infrared spectrometry; iron; metalloenzyme; protein structure function; synthetic enzyme

DESCRIPTION: Research described in this proposal will be aimed at: Understanding how structure influences function and/or the characteristic properties of metalloenzyme active sites. Understanding how protein constraint can influence both the reactivity, and geometric/electronic structure of metalloenzymes. More specifically: Studies will continue to focus on the design and synthesis of analogues for the iron and cobalt sites of nitrile hydratases. Synthetic models, possessing a variety of structures, will be synthesized (using a versatile, one-pot synthesis), structurally characterized (using X-ra crystallography), and then screened for key biophysical properties such as (1) a S=1/2 spin-state (epr, magnetism (both solution and solid state)), and (2) a intense visible band near 700 nm. Reactions between our synthetic models and enzyme substrates (RCN),inhibitor (N3-), and inactivator (NO), will then be monitored both spectrophotometricall and by EPR. By systematically altering the structure of our models, and probing the influence that these changes have on the electronic, and reactivity, properties, we will determine if there is any correlation between structure an reactivity, and determine if spin-state and other electronic properties have a important influence on reactivity. Ligand constraints will be incorporated into our models, as a model for protei constraints, so that we can probe the influence that protein constraints can have on structure and reactivity. For example, by using a ligand with a fixed "backbone" length, angles can be constrained, and, in some cases, opened, so a to create a substrate binding site (entatic state). Similar constraints may be responsible for the reactivity of metalloenzymes. Studies aimed at understanding structure/reactivity relationships in other cysteine-ligated metalloenzymes, such as hydrogenase, and liver alcohol dehydrogenase, will also be pursued.

描述：本提案中描述的研究旨在： 了解结构如何影响功能和/或特征 金属酶活性位点的性质。 了解蛋白质如何 约束可以影响反应性和几何/电子 金属酶的结构。 更具体地说：研究将继续集中于设计和 腈水合酶铁和钴位点类似物的合成。 具有多种结构的综合模型将被合成 （使用多功能的一锅合成），结构表征（使用 X 射线晶体学），然后筛选关键的生物物理特性，例如 作为 (1) S=1/2 自旋态（epr，磁性（溶液和固态））， (2) 700 nm 附近的强烈可见光带。 我们的合成模型与酶底物 (RCN)、抑制剂之间的反应 (N3-) 和灭活剂 (NO)，然后将同时监测 分光光度法和 EPR 法。 通过系统地改变我们模型的结构，并探索 这些变化对电子和反应性的影响， 属性，我们将确定结构之间是否存在任何相关性 反应性，并确定自旋态和其他电子特性 对反应活性有重要影响。 配体约束将被纳入我们的模型中，作为模型 protei 约束，以便我们可以探究蛋白质的影响 限制可能对结构和反应性产生影响。 例如，通过使用 配体具有固定的“主链”长度，角度可以受到限制，并且， 在某些情况下，打开后，会创建一个底物结合位点（惯性状态）。 类似的限制可能是金属酶反应性的原因。 旨在了解其他结构/反应关系的研究 半胱氨酸连接的金属酶，例如氢化酶和肝醇 脱氢酶，也将被追求。