Structural and functional basis of bacterial transcriptional regulation

项目摘要/摘要 抗生素耐药性构成了一个日益普遍的公共卫生挑战，但在 新药的开发没有取得相应的进展。一个重要的障碍在于太多 细菌中的基本生化过程仍然知之甚少。建议的主要目标是 研究旨在从结构上阐明细菌如何响应和适应其环境中的变化 和机械论的立场。使用遗传易驯化的革兰氏阳性菌枯草杆菌作为我们的模型 系统，我们将集中精力阐明铜依赖转录的分子基础。 调节和揭示环境应激反应中酶特异性的驱动因素。铜是 细菌生存所必需的，但过量的这种过渡金属可能是有毒的。因此，铜的水平 必须小心控制。铜出口发生的机制已经被广泛研究， 但人们对铜进口的了解相对较少。我们假设铜的摄取是由铜依赖调节的 转录抑制因子及其控制下的蛋白质。在这里，我们将重点关注一组与之相关的蛋白质 在这些过程中，在详细的结构、分子和细胞水平上研究它们，以更好地了解 它们调节铜的吸收。同时，我们将调查是什么原因导致了一个非常密切相关的激酶家族 具体规范不同的职能。许多细菌种类，包括枯草芽孢杆菌和一些 致病菌株编码一个或多个含有被称为Bergerat折叠的结构域的激酶。尽管他们 结构上的共性，具有这个结构域的酶表现出强烈的生理结合偏好 合伙人。然而，这种特异性是如何被赋予的分子基础还有待研究。我们 假设Bergerat折叠内的变异会影响底物的特异性和动力学，并且可以 以小分子抑制剂为靶点。通过整合结构生物学、微生物学、生物化学、 和计算研究，我们将准备揭示转录控制的新范式。完成度 将促进我们未来对阐明致病的相似途径的研究。 菌株，在细菌系统发育中发现这些家族的新成员，并设计小分子 抑制剂。

Project Summary / Abstract Antibiotic resistance poses an increasingly prevalent public health challenge, but advances in the development of new drugs has not progressed commensurately. A significant impediment lies in that many fundamental biochemical processes in bacteria remain poorly understood. The primary goal of the proposed research is to illuminate how bacteria respond and adapt to changes within their environments from a structural and mechanistic standpoint. Using the genetically tractable Gram positive organism Bacillus subtilis as our model system, we will focus our efforts on elucidating the molecular basis for copper-dependent transcriptional regulation and uncovering the drivers of enzyme specificity during the environmental stress response. Copper is required for bacterial survival, but excess levels of this transition metal can be toxic. Therefore, copper levels must be carefully controlled. The mechanisms by which copper export occurs have been extensively studied, but relatively little is known about copper import. We hypothesize that Cu uptake is regulated by Cu-dependent transcriptional repressors and the proteins under their control. Here, we will focus on a suite of proteins implicated in these processes, studying them at a detailed structural, molecular, and cellular level to better understand how they regulate copper uptake. In parallel, we will investigate what causes a family of very closely related kinases to specifically regulate distinct functions. Many bacterial species, including Bacillus subtilis and a number of pathogenic strains, encode one or more kinases that contain a domain termed the Bergerat fold. Despite their structural commonalities, enzymes with this domain exhibit strong preferences for their physiological binding partners. However, the molecular basis for how such specificity is conferred has yet to be investigated. We hypothesize that variations within the Bergerat fold influence substrate specificity and kinetics and can be targeted by small molecule inhibitors. By integrating tools from structural biology, microbiology, biochemistry, and computational studies, we will be poised to reveal new paradigms in transcriptional control. The completion of the proposed studies will catalyze our future investigations into elucidating analogous pathways in pathogenic strains, uncovering novel members of these families in bacterial phylogeny, and designing small molecule inhibitors.