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.

项目概要/摘要 IGP Synthase (IGPS) 是一个有前途的新型抗生素药物靶点。这种酶起着关键作用 存在于细菌中的色氨酸生物合成中，但不存在于人类中。本提案的总体目标 旨在获得对各种底物-IGPS相互作用的新认识并开发新的IGPS抑制剂 可能具有作为抗感染剂的潜力。因此，这项工作对于理解和 治疗顽固且广泛的耐药细菌。 我们将使用来自结核分枝杆菌的 IGPS (MtIGPS) 作为模型系统。新型 MtIPGS 底物 类似物将以这样的方式进行战略性设计，使得它们相对于酶的行为将 回答有关 IGPS 结构功能关系和催化的问题。一系列新型基材 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 系统地与使用天然底物和非反应性还原剂获得的结果进行比较 底物类似物。这些实验将确定底物功能的作用和重要性 组并揭示哪些必须保留，哪些可以修改为抑制剂 优化。 怀疑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提供对 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 IGPS 配体和活性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。 普遍接受的 IGPS 催化机制始于吸热分子内反应 底物的环化和脱芳构化。 MtIGPS 可能通过强结合或 围绕所得中间体或结构相似的先前转变发生构象塌陷 状态。新型 MtIGPS 抑制剂旨在通过酶促转化为过渡态类似物，无需 转化形成产品将被合成。这些新药物的抑制特性和作用 IGPS 动力学途径上的化合物将被确定。

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