Investigation of Substrate Specificity, Mechanism, and Inhibition of IGPS

IGPS 的底物特异性、机制和抑制研究

• 批准号: 9441461
• 负责人: NINA M GOODEY
• 金额: $ 30.18万
• 依托单位: MONTCLAIR STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-01 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9441461
• 关键词: Active Sites; Adverse effects; Affect; Anabolism; Anti-Bacterial Agents; Anti-Infective Agents; Antibiotics; Area; Bacteria; Behavior; Binding; Biological Models; Biomedical Research; Catalysis; Complement; Complex; Cyclization; Data; Development; Drug Targeting; Drug resistance; Enzymes; Equilibrium; Goals; Growth; Human; Indoles; Investigation; Kinetics; Knowledge; Laboratories; Learning; Ligand Binding; Ligands; Mammals; Metabolic Pathway; Methods; Modification; Molecular Conformation; Mutagenesis; Mutation; Mycobacterium tuberculosis; Pathway interactions; Pharmaceutical Preparations; Pharmacologic Substance; Play; Process; Property; Protein Engineering; Proteins; Reaction; Research; Role; Series; Structure; Structure-Activity Relationship; Substrate Specificity; System; Tryptophan; Work; alpha-glycerophosphoric acid; analog; bacterial resistance; base; catalyst; design; experimental study; functional group; inhibitor/antagonist; innovation; inorganic phosphate; insight; intermolecular interaction; mutant; novel; pathogen; scaffold

Project Summary / Abstract IGP Synthase (IGPS) is a promising new antibiotic drug target. This enzyme plays a critical role in tryptophan biosynthesis in bacteria but is not present in humans. The overall goals of this proposal are to gain new understanding of various substrate-IGPS interactions and to develop new IGPS inhibitors which may have potential as anti-infective agents. Thus, this work is highly relevant to understanding and treating intractable and widespread resistant bacteria. We will use IGPS from M. tuberculosis (MtIGPS) as a model system. New MtIPGS substrate analogs will be strategically designed in such a way that their behavior with respect to the enzyme will answer questions about IGPS structure-function relationships and catalysis. A series of new substrate analogs with altered or removed functional groups and aromatic ring substitutions will be synthesized. The analogs' ability to bind and act as substrates for MtIGPS will be determined. These data will be systematically compared to those obtained with the natural substrate and an unreactive reduced substrate analog. These experiments will determine the role and importance of the substrate functional groups and reveal which ones must be preserved and which ones can be modified for inhibitor optimization. Several MtIGPS active site residues suspected to play a role in ligand binding will be replaced using mutagenesis. The effects of these mutations on kinetic parameters will provide insights into the role of each residue in binding and catalysis. These data will reveal the importance of specific intermolecular interactions between IGPS ligands and the active site, and they complement the data obtained with substrate analogs. The generally-accepted mechanism for IGPS catalysis begins with an endothermic intramolecular cyclization and de-aromatization of the substrate. MtIGPS likely facilitates this step by binding strongly or conformationally collapsing around the resulting intermediate or a structurally-similar preceding transition state. New MtIGPS inhibitors designed to be converted enzymatically into transition state analogs without turning over to form products will be synthesized. The inhibitory properties and the effects of these new compounds on the IGPS kinetic pathway will be determined.

项目摘要 /摘要 IGP合酶（IGP）是一种有希望的新抗生素药物靶标。这种酶起着至关重要的作用 在细菌的色氨酸生物合成中，但人类不存在。该提议的总体目标 是为了获得对各种底物-IGP相互作用的新理解，并开发新的IGP抑制剂 可能具有抗感染剂的潜力。因此，这项工作与理解和 治疗顽固性和广泛的抗性细菌。 我们将使用结核分枝杆菌（MTIGP）的IGP作为模型系统。新的mTIPGS基材 类似物将以战略性设计，以使其相对于酶的行为将会 回答有关IGP结构功能关系和催化的问题。一系列新的基板 将合成具有改变或去除的官能团和芳环取代的类似物。 将确定类似物结合并充当MTIGPS底物的能力。这些数据将是 与自然底物获得的和无反应性降低的系统相比 底物类似物。这些实验将确定底物功能的作用和重要性 小组并揭示必须保留哪些，并且可以修改哪些抑制剂 优化。 涉嫌在配体结合中发挥作用的几个MTIGPS活性位点残基将被替换 使用诱变。这些突变对动力学参数的影响将为您提供见解 每个残基在结合和催化中的作用。这些数据将揭示特定的重要性 IGP配体与活动位点之间的分子间相互作用，它们补充了数据 用底物类似物获得。 IGPS催化的一般接受的机制始于吸热分子内分子 底物的环化和脱染色。 mtigps可能通过牢​​固约束或 在所得的中间体或结构相似的前过渡周围构象折叠 状态。新的MTIGPS抑制剂旨在将酶转化为过渡状态类似物，而无需 将合成成型产品。这些新的抑制特性和影响 将确定IGPS动力学途径上的化合物。

项目成果

Mutational analysis confirms the presence of distal inhibitor-selectivity determining residues in B. stearothermophilus dihydrofolate reductase.

突变分析证实嗜热脂肪芽孢杆菌二氢叶酸还原酶中存在远端抑制剂选择性决定残基。

• DOI: 10.1016/j.abb.2020.108545
• 发表时间: 2020
• 期刊: Archives of biochemistry and biophysics
• 影响因子: 3.9
• 作者: Eck,Tyler;Patel,Seema;Candela,Thomas;LeonH,Katherine;Little,Michael;Reis,NataliaE;Liyanagunawardana,Uththara;Gubler,Ueli;Janson,CherylA;Catalano,Jaclyn;Goodey,NinaM
• 通讯作者: Goodey,NinaM

Insights into the catalytic mechanism of M. tuberculosis indole-3-glycerol phosphate synthase.

深入了解结核分枝杆菌吲哚-3-甘油磷酸合酶的催化机制。

• 发表时间: 2022
• 期刊: FASEB journal : official publication of the Federation of American Societies for Experimental Biology
• 作者: Goodey,NinaM;Cho,Sarah;Maryum,Bhatti;Booter,Huma;Moran,Cintya;Jefferson,Natalie;VanDenBerg,Savannah;Marin,Patryjca;Peralta,Ashley;Margolin,Katherine;Konas,David
• 通讯作者: Konas,David