Mechanical Regulation of Ultra-Sensitivity in E. Coli Flagellar Motors

项目摘要 许多活动细菌物种都表现出集群运动性，这与快速入侵有关 尿路感染（UTIs）的病原体。每年，UTI仅在美国就导致数千人死亡 并对公共卫生系统造成了很大的负担。群集运动与底物相关， 是由细菌鞭毛马达驱动的，鞭毛马达旋转细胞外的螺旋状细丝，对细胞产生推力- 身体虽然趋化性不是群集所必需的，但分子开关的功能使得 电机旋转方向的反转是必不可少的。这个开关是由CheY-P激活的，CheY-P是一种细胞内 由趋化性网络调节的反应调节器。在CheY-P结合后，合作 多子单元开关复合体内的相互作用驱动从逆时针（CCW） 顺时针（CW）构象的可能性越来越大，导致旋转方向的变化。我们 最近的结果表明鞭毛马达感受机械力，其产生于与固体基质的接触， 这导致了开关的抑制。在很短的时间内，电机适应这些力并恢复到正常状态。 能够扭转方向。然而，负责适应的分子基础仍然不清楚。 因此，迫切需要确定开关如何适应机械刺激以促进群集。 如果没有这样的知识，利用抗病毒策略作为治疗方法的潜力， 对抗虫群介导的宿主入侵和抗生素耐药性可能仍然有限。我们的长期目标 目的是促进新的临床有用的抗病毒策略的发展， 集群和殖民。我们在这项申请中的总体目标是确定分子机制 由此开关完美地适应机械信号并促进群集。我们的核心假设是 电机-机械传感（机械信号的传感）导致超灵敏度的调谐， 调节开关内的变构和协同相互作用。拟议工作的理由是 确定超灵敏度的机械控制机制可能会提供一个概念性的 框架的发展战略，以干扰开关适应，并减轻蜂拥而至。在 在完成拟议的研究，这是我们的期望，定量解释的机制， 潜在的开关适应和机械力对超灵敏度的调制。结果预计将 具有重要的积极影响，因为对基板附近开关的详细了解将提供 为生物医学设备中的新型基底设计奠定了坚实的基础，包括导管，这将针对 电动机开关，以抑制蜂拥。

PROJECT ABSTRACT Swarming motility, exhibited by many motile species of bacteria, has been implicated in the rapid invasion of hosts during urinary tract infections (UTIs). Annually, UTIs result in several thousand deaths in the US alone and represent a significant load on the public healthcare system. Swarming motility is substrate-associated and is driven by bacterial flagellar motors that rotate extracellular, helical filaments to generate thrust on the cell- body. Although chemotaxis is not required for swarming, the functioning of a molecular switch that enables reversals in the direction of motor-rotation is indispensable. The switch is activated by CheY-P, an intracellular response-regulator that is regulated by the chemotaxis network. Upon CheY-P-binding, cooperative interactions within the multi-subunit switch-complex drive concerted transitions from counterclockwise (CCW) to clockwise (CW) conformations with increasing likelihood, resulting in changes in the direction of rotation. Our recent results indicate that flagellar motors sense mechanical forces, arising from contact with solid substrates, and that leads to the inhibition of switching. In a short time the motor adapts to these forces and recovers the ability to reverse directions. However, the molecular underpinnings responsible for adaptation remain unclear. Thus, there is a critical need to determine how the switch adapts to mechanical stimuli to promote swarming. Without such knowledge, the potential to capitalize on antivirulence strategies as therapeutic approaches to combat swarming-mediated host-invasion and antibiotic resistance will likely remain limited. Our long-term goal is to contribute toward the development of new clinically useful antivirulence strategies that target bacterial swarming and colonization. Our overall objective in this application is to determine the molecular mechanisms whereby the switch adapts perfectly to mechanical signals and promotes swarming. Our central hypothesis is that motor-mechanosensing (sensing of mechanical signals) results in the tuning of ultra-sensitivity through the modulation of allosteric and cooperative interactions within the switch. The rationale for the proposed work is that a determination of the mechanism of mechanical control of ultra-sensitivity is likely to provide a conceptual framework for the development of strategies to interfere with switch adaptation, and to mitigate swarming. At the completion of the proposed research, it is our expectation to have quantitatively explained the mechanisms underlying switch-adaptation and modulation of ultra-sensitivity by mechanical forces. Results are expected to have an important positive impact because a detailed understanding of switching near substrates will provide a strong foundation for novel substrate-design in biomedical devices, including catheters, which will target the motor-switch to inhibit swarming.