Inter-Bacterial Competition Systems in Burkholderia pseudomallei

描述（由申请方提供）：类鼻疽伯克霍尔德菌是类鼻疽病的病原体，是一种潜在的生物恐怖主义威胁，对大多数常用抗生素具有内在耐药性，这表明对替代抗菌药物的明确需求，以及对这种新兴微生物所采用的致病策略的更好理解。在大肠杆菌中，细菌间的接触依赖性抑制（CDI）系统，包括两个合作伙伴的分泌蛋白被确定为抑制靶细菌的等位基因特异性的方式。由于位于CdiA外蛋白C末端的核酸酶活性而产生的毒性可通过直接结合系统的同源免疫蛋白CdiI来阻断。在B的基因组中发现了该系统的同源物。类鼻疽和相关的非病原体B。泰国茶然而，伯克霍尔德氏菌和E. coliCDI系统，这些系统在伯克霍尔德氏菌生物学中的作用尚不清楚。此外，虽然已经提出了CDI的模型，但CdiA/BtpA蛋白的定位和拓扑结构是未知的，这妨碍了对CDI机制的完全理解。初步数据表明，B.与大肠杆菌素E5 tRNase具有序列相似性的类鼻疽杆菌CdiA同源物BtpA在几种细菌中产生时是有毒的，除非也产生系统的同源免疫蛋白BtpI。为了实现理解这些系统在B生命周期中的作用的长期目标。对于类鼻疽杆菌，本申请将检验BtpA是表面定位的蛋白质的假设，其作为种内和/或种间核酸酶起作用以介导细菌间抑制。目标1中的研究将确定BtpA蛋白的拓扑结构和加工，目标2中的实验将确定BtpA毒性的机制并定义最小抑制单位。最后，目标3中提出的研究将评估BtpA对其他细菌病原体的毒性，并检查其介导种内和/或种间细菌抑制的能力。拟议的研究将有助于了解一种新兴的病原体，并通过研究一类新型细胞毒性蛋白，满足追求新型抗菌疗法的基本需求。

DESCRIPTION (provided by applicant): Burkholderia pseudomallei, the causative agent of melioidosis and a potential bioterrorism threat, is intrinsically resistant to most commonly-prescribed antibiotics, demonstrating a clear requirement for alternative antimicrobials, as well as a better understanding of the pathogenic strategies utilized by this emerging organism. In Escherichia coli, an inter-bacterial contact-dependent inhibition (CDI) system comprised of two-partner secreted proteins was identified that inhibits target bacteria in an allele-specific manner. Toxicity, which is due to nuclease activity located at the C-terminal end of the CdiA exoprotein, is blocked by direct binding of the system's cognate immunity protein, CdiI. Homologs of this system are found in the genomes of B. pseudomallei and the related non-pathogen B. thailandensis. However, distinct differences exist between the Burkholderia and E. coli CDI systems and the role of these systems in Burkholderia biology is not known. Moreover, while a model for CDI has been suggested, the localization and topology of the CdiA/BtpA proteins are unknown, preventing a complete understanding of the CDI mechanism. Preliminary data indicate that the C-terminal region of the B. pseudomallei CdiA homolog, BtpA, which shares sequence similarity with colicin E5 tRNase, is toxic when produced in several bacterial species unless the system's cognate immunity protein, BtpI, is also produced. Toward the long-term objective of understanding the role of these systems in the lifecycle of B. pseudomallei, this application will test the hypothesis that BtpA is a surface-localized protein that functions as an intra- and/or inter-species nuclease to mediate inter-bacterial inhibition. Studies in Aim 1 will determine the topology and processing of the BtpA protein and experiments in Aim 2 will identify the mechanism of BtpA toxicity and define the minimal inhibitory unit. Finally, the research proposed in Aim 3 will assess the toxicity of BtpA to other bacterial pathogens and examine its ability to mediate intra- and/or inter- species bacterial inhibition. The proposed research will contribute to the understanding of an emerging pathogen and, by investigating a novel class of cytotoxic proteins, fulfills an essential need in the pursuit of novel antimicrobial therapies.