Heme-Dependent Chemistry in Aromatic Oxidation

芳香族氧化中血红素依赖性化学

基本信息

批准号：
10819008
负责人：
Aimin Liu
金额：
$ 6.53万
依托单位：
UNIVERSITY OF TEXAS SAN ANTONIO
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-08-01 至 2026-08-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10819008
关键词：
Acids Affect Amino Acids Anabolism Antibiotics Antineoplastic Agents Aromatic Compounds Binding Biochemistry Biogenesis Biological Biological Process Catabolism Catalysis Cations Chemicals Chemistry Crystallography Development Dioxygenases Distal Drug Design Electron Spin Resonance Spectroscopy Environment Enzymes Family Funding Generations Genetic Goals Health Heme Hemeproteins Histidine Human Hydrogen Peroxide Hydroxylation In Situ Isotope Labeling Kinetics Knowledge Lead Learning Ligation Metabolism Mixed Function Oxygenases Molecular Monitor Natural Products Outcome Oxidants Oxidation-Reduction Oxygen Oxygenases Parents Pathway interactions Pharmacologic Substance Play Porphyrins Process Property Protein Engineering Proteins Quinones Reaction Regulation Research Project Grants Roentgen Rays Role Rotation Site Source Specificity Spectrum Analysis Structure System Temperature Testing Time Tryptophan Tyrosine Visit Work analog anti-cancer antimicrobial drug base catalyst chemical reaction cofactor design dopaquinone drug development drug discovery enzyme mechanism heme a insight member millisecond natural antimicrobial novel oxidation pyrroline recruit scaffold secondary metabolite structural determinants thioether x-ray free-electron laser

项目摘要

Abstract of the Parent Research Project Heme-containing enzymes are ubiquitous, contributing to various processes and catalyzing a vast array of oxidative chemistries. Histidine-ligated heme enzymes, such as the heme-dependent aromatic oxygenase (HDAO) superfamily, can use either molecular oxygen or hydrogen peroxide to oxidize their substrates. What is currently not understood are the molecular factors that engender these enzymes with their respective specificities, i.e., after generation of the critical active oxidant, how is a single reaction type catalyzed? The answer to this question will broaden our fundamental understanding of enzyme catalysis, de novo enzyme design, and protein engineering. This application focuses on the structural and mechanistic characterization of three aromatic-oxidizing enzymes of the HDAO superfamily involved in natural product and antimicrobial drug biosynthesis, including antibiotics with a pyrroline moiety or a core quinone structure. These enzymes play essential roles in the biosynthetic pathway of secondary metabolites found in many biologically active natural products and pharmaceutical lead compounds. Our primary goal is to understand the key factors that govern their hydroxylation mechanisms and their differing strategies to form oxidizing intermediates with these enzymes. We have utilized a broad spectrum of approaches to probe the intricate molecular details of these enzyme mechanisms using noncanonical amino acids, substrate analogs, time-resolved UV-vis and EPR spectroscopies, kinetics, and isotope-labeling studies, and we have been successful in trapping on-pathway reaction intermediates in crystallo. The proposed work will test our hypotheses regarding i) how the heme-based oxidant is generated, ii) how oxygen is directed to the substrate, and iii) unravel the structural factors that affect catalysis. The outcome of the proposed studies is an in-depth understanding of these three related catalytic systems to aid the development of scaffolds for rational drug design and discovery processes.

家长研究项目摘要含血红素的酶无处不在，有助于各种过程并催化大量的酶氧化化学。组氨酸连接的血红素酶，例如血红素依赖性芳香酶加氧酶（HDAO）超家族，可以使用分子氧或过氧化氢来氧化其基材。目前尚不了解的是产生这些酶的分子因素它们各自的特殊性，即在产生关键活性氧化剂后，单个反应如何类型催化？这个问题的答案将拓宽我们对酶的基本理解催化、从头酶设计和蛋白质工程。该应用程序侧重于结构 HDAO 超家族中三种芳香族氧化酶的作用机制及其表征天然产物和抗菌药物生物合成，包括带有吡咯啉部分或核心醌结构。这些酶在次级代谢产物的生物合成途径中发挥着重要作用。在许多生物活性天然产物和药物先导化合物中发现的代谢物。我们的主要目标是了解控制其羟基化机制的关键因素及其用这些酶形成氧化中间体的不同策略。我们利用了广泛的使用一系列方法来探测这些酶机制的复杂分子细节非规范氨基酸、底物类似物、时间分辨紫外可见光谱和 EPR 光谱、动力学、和同位素标记研究，我们已经成功捕获途径反应晶体中间体。拟议的工作将测试我们的假设：i）基于血红素的氧化剂的产生，ii) 氧气如何被引导至基材，以及 iii) 揭示产生氧化剂的结构因素影响催化作用。拟议研究的结果是对这三个相关的深入了解催化系统有助于开发合理的药物设计和发现过程的支架。