Structural and Functional Analyses of Toxin-Antitoxin Protein Complexes From Bact

描述(申请人提供)：细菌已经进化出复杂的策略来在他们的环境中竞争和交流。最近在大肠杆菌中发现了一种新的细菌间竞争机制，称为接触依赖生长抑制(CDI)。CDI系统广泛存在于各种革兰氏阴性细菌中，包括几种重要的人类病原体。CDI是由CDIB/CDIA双伙伴分泌系统介导的。CDIB是一种预测的外膜蛋白，它是CDIA外蛋白出口和组装到细胞表面所必需的。CDIA的C-末端结构域(CDIA-CT)具有生长抑制活性，可能被切割并转位到靶细胞胞浆中抑制生长。CDI系统还编码CDI免疫蛋白，结合并灭活CDIA-CT毒素，从而保护CDI+细胞免受自身抑制。值得注意的是，CDIA-CT结构域是多态的，到目前为止已经识别出60多个不同的毒素序列。因此，相应的CdiI蛋白也是高度可变的。这种序列多样性表明CDI系统部署了各种各样的有毒活动。事实上，我们已经发现CDIA-CT结构域表现出许多不同的核酸酶活性。由于CDI免疫蛋白是其同源CDIA-CTs所特有的，每个毒素-免疫复合体的相互作用可能是独一无二的。目前还没有任何CDIA-CT/CDI复合体的结构信息，因此这些蛋白质-蛋白质相互作用的细节完全未知。此外，CdiI蛋白中和CDIA-CT活性的机制尚不清楚。我们建议进行结构和功能分析，以深入了解由细菌CDI系统编码的复杂的毒素免疫网络，这是一个独特的机会，可以阐明当毒素免疫对在进化过程中分化时，特异性结合是如何保持的。 与公共健康相关：细菌已经进化出复杂的策略，在它们的环境中竞争和交流。最近在多种革兰氏阴性细菌中发现了一种新的细菌间竞争机制，称为接触依赖性生长抑制(CDI)。这项建议利用结构/功能分析来获得对CDI系统编码的复杂的毒素免疫蛋白质网络的机械性见解。这项研究将对我们理解细菌的发病机制和生态学产生重大影响，并可能导致新的抗菌疗法的发展。

DESCRIPTION (provided by applicant): Bacteria have evolved complex strategies to compete and communicate in their environments. A new mechanism of inter-bacterial competition, termed contact- dependent growth inhibition (CDI) was recently discovered in Escherichia coli. CDI systems are found in a wide variety of gram-negative bacteria, including several important human pathogens. CDI is mediated by the CdiB/CdiA two-partner secretion system. CdiB is a predicted outer membrane protein that is required for the export and assembly of the CdiA exoprotein onto the cell surface. The C-terminal domain of CdiA (CdiA-CT) contains the growth inhibition activity and is presumably cleaved and translocated into the target cell cytoplasm to inhibit growth. CDI systems also encode CdiI immunity proteins, which bind and inactivate CdiA-CT toxins, thereby protecting CDI+ cells from autoinhibition. Remarkably, the CdiA-CT domain is polymorphic, with well over 60 different toxin sequences identified to date. Accordingly, the corresponding CdiI proteins are also highly variable. This sequence diversity suggests that CDI systems deploy a wide variety of toxic activities. Indeed, we have discovered that CdiA- CT domains exhibit a number of distinct nuclease activities. Because CdiI immunity proteins are specific for their cognate CdiA-CTs, the interactions underlying each toxin- immunity complex are presumably unique. There is currently no structural information available for any CdiA-CT/CdiI complex, and therefore the details of these protein- protein interactions are completely unknown. Moreover, the mechanisms by which CdiI proteins neutralize CdiA-CT activities are not understood. We propose structural and functional analyses to gain insights into the intricate toxin-immunity network encoded by bacterial CDI systems, which represents a unique opportunity to elucidate how specific binding is maintained as toxin-immunity pairs diverge through evolution. PUBLIC HEALTH RELEVANCE: Bacteria have evolved complex strategies to compete and communicate in their environments. A new mechanism of inter-bacterial competition, termed contact-dependent growth inhibition (CDI) has recently been discovered in a wide variety of gram-negative bacterial pathogens. This proposal utilizes a structure/function analysis to gain mechanistic insights into the intricate toxin-immunity protein network encoded by CDI systems. This research will have a significant impact on our understanding of bacterial pathogenesis and ecology and could lead to the development of novel antimicrobial therapies.