Energy Transduction Between Membranes

描述(申请人提供)：革兰氏阴性细菌的外膜是某些营养物质扩散到周质空间和随后进入细胞的运输的障碍。为了获得大量、稀缺的重要营养物质，如铁载体和维生素B12，其运输底物具有亚纳米分子亲和力的活性转运蛋白位于外膜。由于外膜缺乏常规能源，铁铁载体和维生素B12通过外膜主动运输到周质空间的能量来自于细胞质膜的质子动力。细胞膜中的TonB-ExbB-ExbD系统似乎可以获得质子动力，并以未知的形式将其传递给外膜活性转运蛋白。我们的长期目标是了解大肠杆菌细胞质和外膜之间依赖TonB的能量转导的机制。TonB基因的突变使许多物种的毒力大大降低。由于铁的可利用性在传染病中的作用，人们普遍认为TonB系统具有开发新抗生素的新靶点的潜力。更重要的是，我们有能力为更广泛地理解能量传导机制做出贡献。目前还没有TonB依赖的能量转导的范例，但根据同源性，至少有三个这样的系统将PMF转换为几乎所有革兰氏阴性细菌的有用工作。除了ExbB/D蛋白外，还有TolQ/R蛋白(OM完整性所必需的)和MoTA/B蛋白(运动所必需的)。因此，随着广泛原理的出现，我们的结果也将对其他能量转导系统的基本知识做出重要贡献。作为特定的目标，我们将解决TonB系统中的三个基本问题：细胞质膜上TonB能量的决定因素是什么，什么形式的能量被传递到外膜转运体，以及转运体如何被TonB带电？与公共卫生相关：几乎所有的革兰氏阴性细菌都使用TonB系统来绕过其外膜对铁获取的限制。了解TonB系统为外膜运输提供能量的分子机制，将有助于未来开发防止铁摄取的新抗生素。病原体获得铁的能力是感染性疾病的重要毒力决定因素。

DESCRIPTION (provided by applicant): The outer membranes of Gram-negative bacteria are barriers to diffusion of certain nutrients into the periplasmic space and subsequent transport into the cell. To acquire large, scarce, important nutrients such as iron-siderophores and vitamin B12, active transporters with sub-nanomolar affinities for their transport substrates are located in the outer membrane. Because the outer membrane lacks a conventional energy source, energy for active transport of iron-siderophores and vitamin B12 across the outer membrane and into the periplasmic space derives from the protonmotive force of the cytoplasmic membrane. The TonB-ExbB-ExbD system in the cytoplasmic membrane appears to harvest protonmotive force and transduce it in an unknown form to the outer membrane active transporters. Our long-term goal is to understand the mechanism of TonB-dependent energy transduction between the cytoplasmic and outer membranes of Escherichia coli. Mutations in tonB render many species significantly less virulent. Due to the role that iron availability plays in infectious disease, it is widely recognized the TonB system has potential as a novel target for the development of new antibiotics. Additional significance arises from our ability to contribute to a broad understanding of energy transduction mechanisms in general. There are currently no paradigms for TonB-dependent energy transduction and yet based on homologies, there are at least three such systems that transduce the pmf into useful work in virtually all Gram-negative bacteria. Besides the ExbB/D proteins, there are the TolQ/R proteins (required for OM integrity) and the MotA/B proteins (required for motility). Our results will therefore also contribute importantly to basic knowledge of the other energy transduction systems as broad principles emerge. As Specific Aims, we will address three fundamental problems in the TonB system: What are the determinants of TonB energization at the cytoplasmic membrane, what form of energy is transduced to the outer membrane transporters, and how does a transporter become energized by TonB? PUBLIC HEALTH RELEVANCE: Virtually all Gram negative bacteria use the TonB system to circumvent the limitations to iron acquisition posed by their outer membranes. Understanding of the molecular mechanism by which the TonB system energizes transport across the outer membranes will allow future development of new antibiotics that prevent iron uptake. The ability of pathogens to acquire iron is an important virulence determinant in infectious disease.