Bacterial and Molecular Determinants of Mycobacterial Impermeability

项目摘要 非复杂结核分枝杆菌感染的典型疗程包括 四种抗生素，持续至少六个月。多层结构的抗渗性。 长期以来，结核病细胞的包膜一直被认为与这种生物体天生的药物敏感性有关。 而外膜被认为是获得抗生素的主要屏障 在肽聚糖或细胞质中的靶标，包膜的其他方面可能起到作用。此外， 与模式生物大肠杆菌和哺乳动物细胞相比，这个领域只有一个 对可以渗透到结核分枝杆菌的化合物的种类有初步的了解。基于 根据已公布的初步数据，PI假设结核分枝杆菌的不渗透性是一种 包膜成分的结果以及目标位置和化学结构 化合物。检验这一假设的一个主要障碍是缺乏高通量工具来识别 控制化合物跨被膜层渗透的细菌和分子因素。在这 建议，私人投资者发展和部署两种互补的方法来定义细菌和 结核分枝杆菌通透性的分子决定因素。他们将首先测试相对重要性 分子把关中的各种结核分枝杆菌因子。接下来，他们将在全球范围内确定M。 导致菌膜不通透性的结核基因。最后，他们会 全面确定与结核分枝杆菌菌膜相关的结构基序 渗透的或缺乏渗透的。这些目标的成功实现将为医学 改善结核分枝杆菌对现有药物和新发现药物的吸收的化学努力 化合物。PI用来实现这些目标的方法很容易移植到物种上 包膜渗透性也是治疗的限制因素，例如非结核分枝杆菌(NTM) 还有革兰氏阴性。

Project Summary The typical course of treatment for uncomplicated Mycobacterium tuberculosis infection comprises four antibiotics and lasts for at least six months. The impermeability of the multi-layered M. tuberculosis cell envelope has long been linked to the organism’s intrinsically-poor drug susceptibility. While the outer ‘myco’ membrane is hypothesized to be the primary barrier to accessing antibiotic targets in the peptidoglycan or cytoplasm, other facets of the envelope likely contribute. Moreover, compared to the model organism Escherichia coli and to mammalian cells, the field has only a rudimentary understanding of the kinds of compounds that can permeate M. tuberculosis. Based on published and preliminary data, the PIs hypothesize that M. tuberculosis impermeability is a consequence of envelope composition as well as the target location and chemical structure of the compound. A major hurdle to testing this hypothesis is the lack of high-throughput tools for identifying bacterial and molecular factors that control compound permeation across envelope layers. In this proposal, The PIs develop and deploy two complementary methods for defining the bacterial and molecular determinants of M. tuberculosis impermeability. They will first test the relative importance of various M. tuberculosis factors in molecule gate-keeping. Next, they will globally determine the M. tuberculosis genes that contribute to mycomembrane impermeability. Finally, they will comprehensively determine the structural motifs associated with M. tuberculosis mycomembrane permeation or lack thereof. Successful completion of these aims will lay the foundation for medicinal chemistry efforts to improve M. tuberculosis uptake of both existing drugs and newly-discovered compounds. The methods that the PIs use to achieve these aims are easily ported to species for which envelope permeability is also treatment-limiting, e.g., non-tuberculous mycobacteria (NTMs) and Gram-negatives.