Nuclear Magnetic Resonance (NMR) studies of antimicrobial peptides in intact cells

抗菌肽(AMPs)目前是人们非常感兴趣的主题，部分原因是它们在我们的免疫系统中发挥着重要作用，部分原因是它们作为新型疗法的潜力，以对抗日益严重的微生物对传统抗生素的耐药性问题。然而，无论是我们对这些分子如何工作的理解，还是我们将它们用作更好治疗的基础的能力，都存在差距。其中一个差距源于这样一个事实，即只在高度简化的系统中研究AMP的详细机制，该系统充其量只能近似AMP所针对的更复杂的细胞膜。微生物细胞膜包含许多额外的成分，这些成分可能会影响AMP的工作方式，并可能有助于AMP在不损害我们自己细胞的水平上杀死微生物。我们打算通过开发使用高分辨率、定量技术(如核磁共振)来研究完整、完整的细菌和哺乳动物细胞中的AMP来弥合这一差距。然后，我们将把我们的结果与模型系统中的结果进行比较，以更好地理解AMP活动是如何被真实细胞中存在的复杂性修改的。我们还将从鳕鱼中鉴定和表征新的AMP。鳕鱼基因组是潜在的新的和有趣的抗菌肽基序的极好来源，因为鳕鱼具有不发达的适应性免疫系统，但实际上是在病原体的海洋中游泳，因此推测需要特别有效的AMP。这项工作的预期意义包括：在揭示AMP机制的核磁共振研究和实际观察到活动的条件之间提供亟需的联系；开发新的方法，使核磁共振能够应用于在整个细胞范围内研究任何膜蛋白；通过确定和表征新型海洋AMP，使AMP群落以及参与鳕鱼和其他鱼类养殖计划的人获得具有重要意义的信息。

Antimicrobial peptides (AMPs) are currently the subject of significant interest, in part due to their important role in our immune systems and in part due to their potential as novel therapeutics to combat the increasingly serious problem of microbial resistance to conventional antibiotics. However, there are gaps in both our understanding of how these molecules work and in our ability to use them as the basis for better treatments. One such gap results from the fact that the detailed mechanism of AMPs has only been studied in highly simplified systems that only approximate, at best, the much more complex cell envelopes against which the AMPs act. Microbe cell envelopes contain a number of additional components that likely affect how AMPs work and probably contribute to the ability of AMPs to kill microbes at levels that don't harm our own cells. We intend to bridge this gap by developing ways to use high resolution, quantitative techniques, like nuclear magnetic resonance (NMR), to investigate AMPs in whole, intact bacteria and mammalian cells. We will then compare our results to the results in the model systems to better understand how AMP actions are modified by the complexity present in real cells. We will also identify and characterize novel AMPs from cod. The cod genome represents an excellent source of potentially new and interesting antimicrobial peptide motifs, since cod have an underdeveloped adaptive immune system, but are literally swimming in a sea of pathogens and so presumably require especially efficacious AMPs. The anticipated significance of the work includes: supplying a badly needed connection between NMR studies that shed light on the mechanism of AMPs and those conditions under which activity is actually observed; developing new methods that enable the application of NMR to the study of any membrane protein in the context of whole cells; making available information of significance to the AMP community and those involved in cod and other fish breeding programs by identifying and characterizing novel marine AMPs.