STRUCTURES INFLUENCE ON REACTIVITY IN METALLOENZYMES

描述：本提案中描述的研究将针对： 了解结构如何影响功能和/或特征 金属酶活性中心的性质。 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：研究将继续侧重于设计和 腈水合酶铁和钴位点类似物的合成。 将合成具有各种结构的合成模型 （使用通用的一锅法合成），结构表征（使用 X-ra晶体学），然后筛选关键生物物理特性，例如 作为（1）S=1/2自旋状态（epr，磁性（溶液和固态））， （2）700 nm附近的强可见光带。 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 电子探针和EPR。 通过系统地改变我们模型的结构， 这些变化对电子，反应性， 性质，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 对反应性有重要影响。 配体约束将被纳入我们的模型，作为一个模型， 蛋白质的限制，这样我们就可以探测蛋白质 约束可能对结构和反应性有影响。 例如通过使用 对于具有固定“骨架”长度的配体，角度可以被限制，并且，在 在某些情况下，打开，从而产生底物结合位点（内结合状态）。 类似的约束可能是负责金属酶的反应性。 研究旨在了解结构/反应性关系，在其他 半胱氨酸连接的金属酶，如氢化酶和肝醇 脱氢酶，也将被追求。

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.