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Heme-Dependent Chemistry in Tyrosine Oxidation

酪氨酸氧化中血红素依赖性化学

基本信息

批准号：
10475429
负责人：
Aimin Liu
金额：
$ 6.11万
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 2022-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10475429
关键词：
Active Sites Amino Acid Sequence Anabolism Antibiotics Bacterial Infections Biological Carbon Catalysis Cations Characteristics Charge Chemicals Chemistry Coupled Coupling Crystallization Cysteine Cytochrome P450 Cytochrome a Data Dioxygenases Distal Dopa Drug Design Electrons Environment Enzymes Funding Goals Grant Health Heme Histidine Human Hydrogen Bonding Hydrogen Peroxide Hydroxylation Investigation Iron Knowledge Laboratories Lead Levodopa Ligands Malignant Neoplasms Mediating Molecular Mononuclear Mycobacterium tuberculosis Natural Products Oxidants Oxides Oxygenases Pathway interactions Peptides Peroxidases Pharmaceutical Preparations Phenols Play Porphyrins Positioning Attribute Process Production Prosthesis Protein Engineering Proteins Protons Reaction Role Scheme Structure Structure-Activity Relationship System Testing Tryptophan Tuberculosis Tyrosine Tyrosine 3-Monooxygenase antimicrobial drug antineoplastic antibiotics antitumor agent base c new cofactor design drug development drug discovery enzyme activity enzyme mechanism heme a insight member metalloenzyme novel novel therapeutics oxidation pathogen pathogenic bacteria small molecule trait

项目摘要

PROJECT DESCRIPTION A distinguishing trait of heme enzymes is that a high-valent iron-oxo species is a common oxidant for mediating a remarkable array of oxidation reactions. However, one conundrum is that each enzyme in general promotes only a specific type of reaction. How the reaction type is determined after the formation of the key oxidant remains an open question whose answers have implications for our fundamental understanding of enzyme catalysis as well as de novo enzyme design and protein engineering. Because tyrosine is an important building block of natural products, this application focuses on a mechanistic characterization of three heme-dependent tyrosine-oxidizing enzymes. Each of these enzymes employs a mononuclear heme cofactor to oxidize its tyrosine-based substrate. Intriguingly, a cytochrome P450 protein, CYP121 from Mycobacterium tuberculosis, catalyzes an unusual oxidative carbon-carbon cross-coupling reaction instead a more common hydroxylation reaction. We found that SfmD is a new member of the tryptophan dioxygenase superfamily that promotes regioselective monooxygenation of a methylated tyrosine substrate. The peroxidase LmbB2 performs a peroxygenase-type of reaction with an axial ligand of histidine instead of cysteine. These enzymes not only catalyze tyrosine-based oxidation reactions but they are also related to antimicrobial drug development. Given the similarities of the heme-based oxidant and the structure of the substrates, the inevitable question arises regarding the governing factors that determine the catalytic activity of these enzymes. In Aim #1, we will determine the mechanistic and structural characteristics of CYP121. Using a battery of spectroscopic and structural approaches coupled with synthetic probes, we will unveil a novel carbon-carbon coupling mechanism mediated by the P450 enzyme. In Aim #2, we will characterize the structure and mechanism of SfmD with emphasis on how the substrate is positioned to the iron-bound oxidant and the capture of catalytic intermediates. We have already identified that this protein is a novel heme-based oxygenase. Aim #3 is focused on studying the peroxidase reaction catalyzed by LmbB2 that is responsible for L-3,4-dihydroxyphenylalanine (L-DOPA) formation through L-tyrosine hydroxylation. We will utilize small-molecule probes to interrogate mechanistic hypotheses. The in-depth analysis of these three related catalytic systems will test our hypothesis regarding how the heme-bound oxidant is generated and directed to the aromatic substrates, unravel the structure-function relationships of the heme enzymes of seemingly unrelated superfamilies at a higher level, and develop underlying mechanisms further aiding rational drug design and discovery processes.

项目描述