Salicylate production in siderophore biosynthesis

铁载体生物合成中水杨酸的产生

基本信息

批准号：
8288743
负责人：
AUDREY L LAMB
金额：
$ 28.81万
依托单位：
UNIVERSITY OF KANSAS LAWRENCE
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-07-15 至 2014-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8288743
关键词：
AIDS chemotherapy Acids Active Sites Adopted Affinity Anabolism Antibiotic Resistance Bacteria Binding Biological Models Burn injury Cancer Patient Catalysis Chelating Agents Chorismate Mutase Crystallography DNA Sequence Rearrangement Dependence Drug Design Electrostatics Environment Enzymatic Biochemistry Enzyme Inhibitor Drugs Enzyme Inhibitors Enzyme Kinetics Enzymes Family Free Energy Goals Growth Health Human Immune Individual Infant Infection Iron Isotopes Kinetics Laboratories Lead Link Lung Lyase Molecular Molecular Conformation Molecular Weight Motion Mutation Pathogenesis Pathway interactions Patients Production Property Protein Family Pseudomonas aeruginosa Publishing Pyruvate Quantum Mechanics Reaction Relative (related person)Research Risk Role Siderophores Solvents Structure Structure-Activity Relationship Testing Thermodynamics Vertebral column Virulence Work antimicrobial drug base cystic fibrosis patients drug discovery enzyme mechanism high throughput screening inhibitor/antagonist innovation insight member molecular dynamics molecular mechanics pathogen peptide synthase pyochelin salicylate small molecule structural biology

项目摘要

DESCRIPTION (provided by applicant): Siderophores are low molecular weight iron chelators produced by bacteria to scavenge iron from the environment, and are frequently required for bacterial virulence. In iron limiting environments such as the human host, if the production of the siderophore is disrupted, the bacteria cannot acquire the iron required for growth and survival. Pseudomonas aeruginosa are notoriously antibiotic-resistant and are able to colonize immuno-compromised patients and the lungs of patients with cystic fibrosis. The long-term goal of the research in this laboratory is to provide a detailed mechanistic and structural understanding of enzymes in the siderophore biosynthetic pathways that are targets for drug discovery. Toward that end, we propose to investigate the nonribosomal peptide synthetase accessory enzymes that produce salicylate, a precursor incorporated into the siderophore pyochelin from the bacteria P. aeruginosa. The two enzymes to be studied, the isochorismate synthase (PchA) and isochorismate-pyruvate lyase (PchB), are ideal model systems for making fundamental advances in understanding the reaction pathways of catalysis. We will use a multifaceted approach that includes enzyme kinetic analysis, structural biology and computational enzymology, and identify inhibitors of the enzymes through high-throughput screening. Our first specific aim will be to determine the relative catalytic contributions of a reactive substrate conformation and electrostatic transition state stabilization in the pericyclic reaction mechanisms performed by the isochorismate- pyruvate lyase that has adventitious chorismate mutase activity. We will define the structure- function relationships of the Mg+2 dependent isochorismate synthase, a member of the MST family of proteins in our second aim. Finally, our third aim is to identify small molecule inhibitors of salicylate production, and determine their mode of action. PUBLIC HEALTH RELEVANCE: Pseudomonas aeruginosa is a dangerous opportunistic pathogen that is a common cause of infections in susceptible hosts, including cancer patients undergoing chemotherapy, AIDS patients, those with immune deficiencies, cystic fibrosis patients, burn patients and other at risk individuals including infants. The goal of this work is to provide a fundamental understanding of two enzymes that promote virulence in P. aeruginosa so that this information can be exploited to generate new antimicrobial drugs.

描述（由申请人提供）：铁载体是细菌产生的低分子量铁螯合剂，用于从环境中清除铁，并且通常是细菌毒力所必需的。在人类宿主等铁限制环境中，如果铁载体的产生被破坏，细菌就无法获得生长和生存所需的铁。众所周知，铜绿假单胞菌对抗生素具有耐药性，能够在免疫功能低下的患者和囊性纤维化患者的肺部定殖。该实验室研究的长期目标是对作为药物发现目标的铁载体生物合成途径中的酶提供详细的机制和结构了解。为此，我们建议研究产生水杨酸的非核糖体肽合成酶辅助酶，水杨酸是一种融入铜绿假单胞菌铁载体绿脓杆菌中的前体。要研究的两种酶，异分支酸合酶 (PchA) 和异分支酸-丙酮酸裂解酶 (PchB)，是在理解催化反应途径方面取得根本进展的理想模型系统。我们将采用多方面的方法，包括酶动力学分析、结构生物学和计算酶学，并通过高通量筛选来鉴定酶的抑制剂。我们的第一个具体目标是确定具有外来分支酸变位酶活性的异分支酸-丙酮酸裂合酶执行的周环反应机制中反应性底物构象和静电过渡态稳定的相对催化贡献。我们的第二个目标是定义 Mg+2 依赖性异分支酸合酶（MST 蛋白家族的成员）的结构-功能关系。最后，我们的第三个目标是鉴定水杨酸产生的小分子抑制剂，并确定它们的作用方式。公共卫生相关性：铜绿假单胞菌是一种危险的机会性病原体，是易感宿主感染的常见原因，包括接受化疗的癌症患者、艾滋病患者、免疫缺陷患者、囊性纤维化患者、烧伤患者和其他高危人群（包括婴儿）。这项工作的目标是提供对两种促进铜绿假单胞菌毒力的酶的基本了解，以便利用这些信息来产生新的抗菌药物。