Dihydropteridine Reductase: Mechanism of Enzyme Action

二氢蝶啶还原酶：酶作用机制

基本信息

批准号：
8011316
负责人：
RUEL Z B DESAMERO
金额：
$ 11.53万
依托单位：
YORK COLLEGE
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-01-01 至 2013-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8011316
关键词：
7,8-dihydrobiopterin Address American Applications Grants Basic Science Binding Biochemical Reaction Biochemistry Brain Caenorhabditis elegans Catalysis Catecholamines Chemicals Chemistry Collaborations Complex Cysteine Defect Department of Defense Development Diffusion Dihydropteridine Reductase Deficiency Disease Dinucleoside Phosphates Disease Disulfides Docking Drug Design Engineering Ensure Enzymatic Biochemistry Enzymes Event Face Fluorescence Funding Future Goals Grant Growth Factor Health Hormone Receptor Human Hydrogen Peroxide Indoles Investigation Kinetics Letters Ligands Measurement Measures Mediating Medicine Metals Molecular Monitor NADH Neurotransmitters Niacinamide Nutrient Oxidants Oxidoreductase Paper Petroleum Play Positioning Attribute Preparation Process Productivity Protein Biochemistry Protein Conformation Proteins Publications Published Comment Quinones Rattus Reaction Recycling Reducing Agents Regulation Reporting Research Research Personnel Research Support Roentgen Rays Role Secure Serotonin Societies Source Spectrum Analysis Structure Substrate Interaction Sulfhydryl Compounds System Techniques Tryptophan Work Writing base career cofactor college dihydropteridine reductase dimer enzyme mechanism enzyme substrate improved inhibitor/antagonist insight instrument interest monomer nervous system disorder programs protein structure simulation small molecule temperature jump tetrahydrobiopterin willingness

项目摘要

DESCRIPTION (provided by applicant): Enzyme-substrate interactions have long been recognized as representing an extreme expression of structural complementarities in biological chemistry. An enzymatic reaction can be broken down into two main aspects. The first involves the diffusion-controlled formation of the encounter complex, and the second involves the appropriate structural and dynamical arrangements of the enzyme domains as dictated by the reaction chemistry. It is the long-term goal of the PIs research program to investigate enzyme-small molecule (inhibitor, substrate, or cofactor) interactions in order to unravel the structural and dynamical aspects of its reaction. This proposal aims to study the hydride transfer mechanism involved in DHPR. Dehydropteridine reductase (DHPR) catalyses the NADH-mediated reduction of quinonoid dihydrobiopterin (qBH2) to yield tetrahydrobiopterin (BH4). BH4 functions as an essential cofactor, and its absence leads to depletion in the brain of precursors of catecholamine and serotonin neurotransmitters. Regulation of DHPR became of interest when a new form of hyperphenylalanineanemia (atypical phenylketonuria) associated with a defect in BH4 recycling was discovered. In the light of other new emerging functions, there is a growing need to completely understand how DHPR works. The overall objective of the investigation is to understand the structural changes that occur when DHPR binds cofactor, substrate or inhibitor. Using advanced fluorescence and vibrational techniques, we will identify catalytically important bonds as well as detect major protein conformational changes that may accompany binding and catalysis. We will also look at the dynamical aspects of the binding and catalytic events using fluorescence-based temperature-jump studies. The results will be married to the solved X-ray crystal structures of DHPR to gain insights into the mechanism of enzyme action. In particular, we would like to address the mechanistic role of the cofactor and the conserved Cys residue. Since DHPR belongs to a superfamily of biologically important short-chain dehydrogenases/reductases (SDRs), it is hoped that the gathered results could provide some mechanistic details useful for studying SDRs. SDRs have diverse functions. Some are good pharmacological targets while others are important in controlling the cellular availability of a hormone receptor ligand. There are also those that have essential role in growth factors or nutrient synthesis. Understanding the enzymology of DHPR at the molecular level will play a part in realizing the untold potential for drug design, enzyme regulation, and enzyme engineering. PUBLIC HEALTH RELEVANCE: Enzyme-substrate interactions have long been recognized as representing an extreme expression of structural complementarities in biological chemistry. Basic research geared towards understanding the inner workings of an enzyme system, like dihydropteridine reductase (DHPR), is important if cures for the diseases caused by a malfunctioning or deficient enzyme are to be found. Regulation of DHPR became of interest with the discovery of atypical phenylketonuria, a neurological disorder, associated with a defect in tetrahydrobiopterin recycling.

描述（由申请人提供）：酶-底物相互作用长期以来被认为代表生物化学中结构互补性的极端表达。酶促反应可以分为两个主要方面。第一个涉及扩散控制的相遇复合物的形成，第二个涉及由反应化学决定的酶结构域的适当结构和动力学排列。 PI 研究计划的长期目标是研究酶与小分子（抑制剂、底物或辅因子）的相互作用，以揭示其反应的结构和动力学方面。本提案旨在研究DHPR中涉及的氢化物转移机制。脱氢蝶啶还原酶 (DHPR) 催化 NADH 介导的醌类二氢生物蝶呤 (qBH2) 还原，生成四氢生物蝶呤 (BH4)。 BH4 是一种重要的辅助因子，它的缺失会导致大脑中儿茶酚胺和血清素神经递质的前体消耗。当发现与 BH4 回收缺陷相关的一种新形式的高苯丙氨酸血症（非典型苯丙酮尿症）时，DHPR 的调节引起了人们的兴趣。鉴于其他新出现的功能，越来越需要完全了解 DHPR 的工作原理。研究的总体目标是了解 DHPR 结合辅因子、底物或抑制剂时发生的结构变化。使用先进的荧光和振动技术，我们将识别具有催化作用的重要键，并检测可能伴随结合和催化的主要蛋白质构象变化。我们还将使用基于荧光的温度跳跃研究来研究结合和催化事件的动力学方面。结果将与已解析的 DHPR 的 X 射线晶体结构结合起来，以深入了解酶的作用机制。特别是，我们想解决辅因子和保守的半胱氨酸残基的机制作用。由于 DHPR 属于生物学上重要的短链脱氢酶/还原酶 (SDR) 超家族，因此希望收集的结果能够为研究 SDR 提供一些有用的机制细节。 SDR 具有多种功能。有些是良好的药理学靶点，而另一些则在控制激素受体配体的细胞可用性方面很重要。还有一些在生长因子或营养合成中发挥重要作用的物质。在分子水平上了解 DHPR 的酶学将有助于实现药物设计、酶调节和酶工程的巨大潜力。公共卫生相关性：长期以来，酶-底物相互作用一直被认为代表了生物化学中结构互补性的极端表达。如果要找到治疗因酶功能障碍或缺陷引起的疾病的方法，旨在了解二氢蝶啶还原酶 (DHPR) 等酶系统内部运作的基础研究非常重要。随着非典型苯丙酮尿症（一种与四氢生物蝶呤回收缺陷相关的神经系统疾病）的发现，DHPR 的调节引起了人们的兴趣。