PROTEIN STRUCTURE AND FUNCTION

我们正在研究甲酰四氢叶酸合成酶，以检测一种 酶学者感兴趣的问题的数量；机理，底物 小配体对结构的结合和调控。概述了这些实验 旨在增加我们对两国关系的理解 该酶的结构和催化活性，从而增加我们的 对酶的一般了解。 我们计划使用~(13)C、~1H和~(31)P核磁共振弛豫技术来研究配体 有约束力的。这些研究将在不同的条件下进行，以确定 核与酶结合的锰(II)之间的核间距； 单体和单体的结合部位存在可检测的差异 四聚体，以及配基是否发生构象变化 有约束力的。将使用紫外差分和荧光光谱来研究 三磷酸腺苷和三磷酸腺苷类似物与酶的相互作用。各种实验都是 建议寻找酶结合甲酰基存在的证据 作为中间体的磷酸盐。三磷酸腺苷的结合部位将由 使用紫外光诱导的可光激活的ATP类似物的交联，随后 蛋白质分解消化。阳离子诱导单体重缔合反应的研究 并提出了反应的建议。装配机制将遵循以下步骤 紫外差分光谱和常规和快速动力学技术。 在一些实验中，我们将对梭状芽胞杆菌与 酶，正在研究的主要蛋白质，以及酵母酶。主修 原核酶和真核酶之间存在结构差异， 这就形成了这些利益的比较。

We are studying the enzyme, formyltetrahydrofolate synthetase, to examine a number of problems of interest to enzymologists; mechanism, substrate binding and control of stucture by small ligands. The experiments outlined are designed to increase our understanding of the relationship between structure and catalytic activity of this enzyme and thereby add to our understanding of enzymes in general. We plan to use 13C, 1H and 31P NMR relaxation techniques to study ligand binding. The studies will be done under different conditions to determine internuclear distances between the nucleus and enzyme-bound Mn(II); whether detectable differences exist in the binding sites of the monomer and tetramer, and whether conformational changes are produced upon ligand binding. UV-difference and fluorescence spectroscopy will be used to study ATP and ATP-analog interactions with the enzyme. Various experiments are proposed to find evidence for the existence of an enzyme-bound formyl phosphate as an intermediate. The binding site for ATP will be explored by using UV-induced crosslinking of photo-activatable ATP analogs, followed by proteolytic digestion. Studies of the cation-induced monomer reassociation reaction are also proposed. The assembly mechanism will be followed by UV-difference spectroscopy and conventional and rapid kinetic techniques. In some of the experiments comparisons will be made between the clostridial enzyme, the primary protein being studied, and the yeast enzyme. Major structural differences exist between the procaryotic and eucaryotic enzyme, which makes these comparisons of interest.