Mechanisms of Sap-mediated Antimicrobial Peptide Resistance

Sap 介导的抗菌肽耐药机制

• 批准号: 8177320
• 负责人: MARGARET E BAUER
• 金额: $ 23.1万
• 依托单位: INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-30 至 2013-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8177320
• 关键词: ATP Hydrolysis; Anti-Bacterial Agents; Area; Bacteria; Binding; Binding Proteins; Biological Models; Cytoplasm; Data; Defensins; Development; Dipeptides; Disease; Epithelial Cells; Family; Genes; Genital system; Genome; Goals; Gram-Negative Bacteria; HIV; Haemophilus influenzae; Hemophilus ducreyi; Host Defense; Human; Human Volunteers; Immune system; Infection; Invaded; Ion Channel; Learning; Mediating; Membrane; Methodology; Methods; Microbe; Modeling; Nontypable Haemophilus influenza; Oligopeptides; Organism; Outcome Study; Parents; Peptide Transport; Peptides; Periplasmic Binding Proteins; Periplasmic Proteins; Phagocytes; Protein Binding; Proteins; Pump; Reporting; Research; Resistance; Role; Sexually Transmitted Diseases; Skin; Specificity; Study models; Surface; Therapeutic; Therapeutic Agents; Ulcer; Virulence; Virulence Factors; Work; antimicrobial; antimicrobial drug; antimicrobial peptide; base; cathelicidin; combat; hCAP-18 protein; human disease; killings; member; microbial; microbicide; mutant; novel; paralogous gene; pathogen; periplasm; permease; research study; tool; transmission process; uptake

DESCRIPTION (provided by applicant): Antimicrobial peptides (APs), including defensins and cathelicidins, are an important component of the innate immune system for combating bacteria and other microbial invaders. Many Gram-negative pathogens utilize the Sensitive to Antimicrobial Peptides (Sap) transporter to survive attack by APs during infection. The Sap transporter has been shown to confer resistance to a wide variety of APs and contributes significantly to virulence. Thus, the Sap transporter is an excellent candidate for targeting development of novel antimicrobial therapeutics that would likely be effective against many Gram-negative pathogens. However, little is known about how the transporter functions with APs to confer resistance to their killing effects. The long-term goal of this project is to understand the mechanism of action of the Sap transporter so that novel antimicrobial agents can be developed that target this transporter. Using the Sap transporter of Haemophilus ducreyi as a model system, we have begun to study the role of specific Sap proteins in conferring AP resistance. H. ducreyi is the causative agent of chancroid, a sexually transmitted genital ulcer disease that facilitates transmission and acquisition of human immunodeficiency virus (HIV) and contributes to the spread of HIV in areas with endemic chancroid. H. ducreyi is one of few pathogens amenable to human inoculation experiments, so studies of the Sap transporter and its interactions with APs can be directly extended to human disease. Our preliminary data with the Sap transporter of H. ducreyi raise questions about the proposed model of the mechanism of Sap transporter activity, including the role of the periplasmic peptide binding component, SapA, in transporter activity and in defining the specificity of the transporter for certain APs. In this proposal, we will establish methods to directly examine the effects of the Sap transporter on the human cathelicidin AP LL37 and use these methods to determine the role of SapA in transporter activity, define possible additional proteins involved in transporter function, and determine which components of the transporter confer specificity for transport of only certain APs. Outcomes of these studies provide the tools required to study the mechanism of action of the Sap transporter and will define the repertoire of proteins that interact specifically with APs for Sap-mediated transport. Data from this proposal will thus provide the methodologies and preliminary data for an R01 application to study in greater detail the interactions of the Sap transporter with APs, which is necessary for developing antimicrobials that target this important virulence factor. PUBLIC HEALTH RELEVANCE: Many gram-negative bacteria use the Sap transporter to avoid being killed by antimicrobial peptides the body produces as a defense against invading microbes. The conservation of this virulence factor among significant pathogens makes it an attractive candidate for development of new antimicrobial therapies; however, the mechanism of action of this transporter is not well understood. The work outlined in this application will define how the Sap transporter aids the bacteria in survival in the host and will define the proteins involved; information learned from these studies will be important for developing new antimicrobial therapeutics that block the activity of this transporter.

说明(申请人提供)：抗菌肽(AP)，包括防御素和抗菌素，是对抗细菌和其他微生物入侵的天然免疫系统的重要组成部分。许多革兰氏阴性菌在感染过程中利用对抗菌肽(SAP)敏感的转运蛋白在AP的攻击下存活。SAP转运蛋白已被证明对多种AP具有抗性，并对毒力有重要贡献。因此，SAP转运体是靶向开发可能对许多革兰氏阴性病原体有效的新型抗菌疗法的极佳候选者。然而，关于转运蛋白如何与AP一起发挥作用以抵抗其杀戮效果，人们知之甚少。该项目的长期目标是了解SAP转运蛋白的作用机制，以便开发针对该转运蛋白的新型抗菌剂。以杜氏嗜血杆菌的SAP转运蛋白为模型系统，我们已经开始研究特定SAP蛋白在AP抗性中的作用。杜克雷嗜血杆菌是下巴的病原体，这是一种性传播的生殖器溃疡疾病，有助于人类免疫缺陷病毒(HIV)的传播和获得，并导致艾滋病毒在地方性下巴流行地区的传播。杜氏嗜血杆菌是少数几种适合人类接种实验的病原体之一，因此对SAP转运蛋白及其与AP相互作用的研究可以直接扩展到人类疾病。我们对H.ducreyi的SAP转运体的初步数据对所提出的SAP转运体活性机制模型提出了质疑，包括胞外肽结合成分SapA在转运体活性中的作用以及在定义转运体对某些AP的特异性方面的作用。在这项提案中，我们将建立方法来直接检测SAP转运体对人长春新碱AP LL37的影响，并使用这些方法来确定SapA在转运体活性中的作用，定义可能参与转运体功能的额外蛋白，并确定转运体的哪些成分仅对某些AP的转运体具有特异性。这些研究的结果提供了研究SAP转运蛋白作用机制所需的工具，并将确定与SAP介导的转运蛋白特异性相互作用的蛋白质谱系。因此，这项提案的数据将为R01应用提供方法学和初步数据，以更详细地研究SAP转运体与AP的相互作用，这对于开发针对这一重要毒力因子的抗菌剂是必要的。 公共卫生相关性：许多革兰氏阴性细菌使用SAP转运蛋白来避免被人体产生的抗菌肽杀死，以防御入侵的微生物。这种毒力因子在重要病原体中的保守性使其成为开发新的抗菌疗法的候选药物；然而，这种转运蛋白的作用机制尚不清楚。本申请中概述的工作将定义SAP转运蛋白如何帮助细菌在宿主中生存，并将定义涉及的蛋白质；从这些研究中获得的信息将对于开发阻断该转运蛋白活性的新的抗菌疗法将是重要的。