Virulence gene regulators of enteric bacterial pathogens: Determining the structural and functional mechanisms of small molecule and polypeptide inhibitors

肠道细菌病原体的毒力基因调节因子：确定小分子和多肽抑制剂的结构和功能机制

• 批准号: 10586700
• 负责人: Fredrick Jon Kull
• 金额: $ 62.54万
• 依托单位: DARTMOUTH COLLEGE
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-11-17 至 2027-10-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10586700
• 关键词: Acinetobacter; Address; Arabinose; Artificial Intelligence; Belief; Binding; Biochemistry; Biological Models; Categories; Characteristics; Citrobacter rodentium; Collaborations; DNA; DNA Binding; DNA Binding Domain; Dimerization; Enteral; Environment; Escherichia; Family; Family member; Family suidae; Fatty Acids; Gene Expression; Gene Expression Regulation; Genetic Transcription; Genetic study; Goals; Homologous Gene; Human; Klebsiella; Knowledge; Legionella; Ligand Binding; Ligand Binding Domain; Ligands; Machine Learning; Mediating; Metabolism; Microbiology; Molecular; Molecular Conformation; Morbidity - disease rate; Pathogenesis; Pathogenicity; Positioning Attribute; Protein Family; Proteins; Pseudomonas; Publishing; Regulator Genes; Regulon; Research; Resolution; Role; Salmonella; Salmonella enterica; Shigella; Signal Transduction; Specificity; Structure; Testing; Therapeutic; Vibrio; Vibrio cholerae; Virulence; Work; Yersinia; alpha helix; biological adaptation to stress; combat; dimer; enteric pathogen; enteroaggregative Escherichia coli; enterotoxigenic Escherichia coli; human morbidity; human mortality; inhibitor; innovation; long chain fatty acid; member; mortality; multidisciplinary; novel; pathogen; pathogenic bacteria; polypeptide; protein function; response; small molecule; structural biology; trait; transcription factor; virulence gene

Virulence gene regulators of enteric bacterial pathogens: Determining the structural and functional mechanisms of small molecule and polypeptide inhibitors Summary: The AraC/XylS family is one of the largest families of bacterial transcription factors with ~16K members distributed amongst 81% of sequenced bacterial species. Family members are present in pathogenic genera including Acinetobacter, Escherichia, Klebsiella, Legionella, Pseudomonas, Salmonella, Shigella, Vibrio, and Yersinia. The regulon of any given family member typically encompasses one of three categories: metabolism, stress response, or pathogenesis. Those involved in metabolism or stress response often have well characterized ligands. For example, AraC regulates the expression of genes involved in arabinose metabolism and its activity is modulated by arabinose. In contrast, small molecule ligands have not been identified for the vast majority of virulence regulators within the AraC family (hereafter referred to as AraC-VRs), which has led to the commonly held belief that the AraC-VR branch of the family has lost the ability to respond to ligands. Published work by us and others –and our preliminary studies– suggest that this assumption is incorrect. Additionally, a large family of endogenously encoded polypeptides, ANRs for AraC negative regulators, have been discovered that inhibit AraC-VRs though an unknown mechanism. The long-term goal of this project is to define the structural and molecular mechanisms underlying virulence gene regulation. The specific objectives of this proposal are to determine how AraC family members including Rns, a primary virulence regulator in enterotoxigenic E. coli (ETEC), are inhibited by 1) small molecule fatty acids and 2) AraC negative regulators (ANRs). Our central hypothesis is that Rns must dimerize in order to bind to DNA and regulate transcription and that these inhibitors block this by distinct mechanisms. The motivating rationale for these studies is that they will identify the molecular and structural requirements for inhibiting virulence gene expression, and will be tested by three specific aims: 1) Determine the structural mechanism by which ligand binding and dimerization regulates Rns activity; 2) Test our hypothesis that the Rns homolog RegA is regulated in the same manner; 3) Determine the mechanism by which ANRs inhibit Rns activity, and clarify if this is distinct from the inhibitory mechanism of small molecule fatty acids. This project is innovative in that the basic molecular mechanisms by which these proteins are regulated are not understood. Our multidisciplinary team, with expertise in microbiology, biochemistry, and structural biology, is uniquely positioned to undertake the proposed studies to determine these mechanisms. This research is significant, not only because it will answer outstanding questions of how AraC proteins function in enteric pathogens, but because we expect to demonstrate that AraC family proteins from a wide variety of enteric pathogens share a common mechanism of being inhibited by fatty acids. This will open up new possibilities for therapeutic strategies to combat global mortality and morbidity.

肠细菌病原体的毒力基因调节剂：确定结构和功能 小分子和多肽抑制剂的机制 概括： ARAC/Xyls家族是〜16K成员的细菌转录因子的最大家族之一 分布在81％的测序细菌物种中。家庭成员存在于致病属中 包括ACINETOBACTER，ESCHERICHIA，KLEBSIELLA，军团菌，假单胞菌，沙门氏菌，志贺氏菌，颤音，弧菌和 Yersinia。任何给定家庭成员的常客通常包括三类之一：代谢， 压力反应或发病机理。那些参与新陈代谢或压力反应的人通常有很好 特征配体。例如，ARAC调节与阿拉伯糖代谢有关的基因的表达 它的活性由阿拉伯糖调节。相反，尚未确定小分子配体的配体 ARAC家族中的绝大多数病毒调节剂（以下称为ARAC-VRS） 人们普遍认为，家庭的ARAC-VR分支机构失去了对配体反应的能力。 我们和其他人以及我们的初步研究发表的工作表明，这一假设是不正确的。 此外，大型内源编码的多肽，用于ARAC负调节剂的ANR具有 被发现通过未知机制抑制ARAC-VR。 该项目的长期目标是定义病毒的结构和分子机制 基因调节。该提案的具体目标是确定ARAC家庭成员如何包括 RNS是肠毒素大肠杆菌（ETEC）中的主要病毒调节剂，被1）小分子脂肪抑制 酸和2）ARAC负调节剂（ANRS）。我们的中心假设是RN必须二聚 与DNA结合并调节转录，并通过不同的机制阻止了这些抑制剂。这 这些研究的激励理由是，它们将确定分子和结构性要求 抑制病毒基因表达，并将通过三个特定目的进行测试：1）确定结构 配体结合和二聚化调节RNS活性的机制； 2）检验我们的假设 RNS同源性雷加以相同的方式进行调节； 3）确定ANR抑制RN的机制 活性，并确定这是否与小分子脂肪酸的抑制作用不同。 该项目具有创新性，因为调节这些蛋白质的基本分子机制不是 理解。我们的多学科团队，具有微生物学，生物化学和结构生物学方面的专业知识，是 独特的定位以进行拟议的研究以确定这些机制。这项研究是 重要的是，不仅因为它将回答有关ARAC蛋白在Enter中如何发挥作用的杰出问题 病原体，但因为我们期望证明来自各种各样的ARAC家族蛋白 病原体具有被脂肪酸抑制的共同机制。这将为 打击全球死亡率和发病率的治疗策略。