Host-mediated zinc sequestration during Acinetobacter baumannii infection

项目摘要 鲍曼不动杆菌是一种重要的院内致病菌，可引起一系列疾病，包括 呼吸道和泌尿道感染、脑膜炎、心内膜炎、伤口感染和菌血症。事实上，我认为， A.在美国的一些地区，鲍曼不动杆菌现在占所有重症监护室感染的20%。 肺炎是最常见的表现。探讨了A.鲍曼尼哈斯 这种细菌对几乎所有抗生素都能迅速产生耐药性。识别 治疗干预的新靶点对于我们保护公众健康的能力至关重要。 新的传染病威胁。治疗发展的一个有希望的潜在领域涉及靶向 细菌进入营养金属或金属处理。这种策略是基于这样一个事实，即所有细菌 病原体需要营养金属才能在宿主体内定植， 深刻地影响了对感染易感性。宿主蛋白钙卫蛋白（CP）是最重要的蛋白质之一， 免疫介导的金属限制和CP的贡献者通过螯合 锌（Zn）和锰（Mn）。在本申请中，我们描述了我们使用CP作为探针， 发现A.鲍曼不动杆菌参与CP依赖性条件下的生存 锌饥饿。该基因座编码保守的COG 0523 GTP酶家族的一个成员， 命名为β-谷氨酰胺诱导的GTdA（ZigA），和我们命名为D-丙氨酸D-丙氨酸羧肽酶的D-丙氨酸羧肽酶， 锌调节脂蛋白A（ZrIA）。我们已经发现ZigA是一种金属伴侣蛋白， 客户蛋白质，并需要在锌饥饿的条件下释放生物可利用的锌池。 这一发现将ZigA确立为自然界中Zn金属伴侣的第一个例子。此外，我们的模型 预测在Zn胁迫期间需要ZrlA来维持细胞壁结构。基于这些基本的 发现，我们假设，在锌饥饿，A。鲍曼不动杆菌将锌转移到需要锌的关键客户端 蛋白质，同时还激活参与肽聚糖重塑的羧肽酶的表达 其替代了需要Zn的蛋白质的活性的损失。为了验证这一假设，我们建议 一系列旨在了解的机制和病理生理后果的实验， A.肺炎发病过程中鲍曼不动杆菌对饮食和宿主锌剥夺的反应 在这些研究中，我们将（i）定义在CP条件下ZigA Zn-金属伴侣的靶标， （ii）阐明ZrlA在条件下对肽聚糖重塑的贡献 的锌剥夺，和（iii）确定锌分布的重要性，在发病过程中的A。 鲍氏肺炎。这些结果将为A.鲍曼尼回应 营养剥夺的脊椎动物宿主，并奠定了基础，创造肽疗法 基于通过营养金属螯合抑制微生物生长的CP支架。

PROJECT SUMMARY Acinetobacter baumannii is an important nosocomial pathogen that causes a range of diseases, including respiratory and urinary tract infections, meningitis, endocarditis, wound infections, and bacteremia. In fact, A. baumannii is now responsible for up to 20% of all intensive care unit infections in some regions of the world with pneumonia being the most common presentation. The clinical significance of A. baumannii has been propelled by this organism's rapid acquisition of resistance to virtually all antibiotics. The identification of novel targets for therapeutic intervention is critical to our ability to protect the public health from this emerging infectious threat. One promising potential area of therapeutic development involves targeting bacterial access to nutrient metal or metal handling. This strategy is based on the fact that all bacterial pathogens require nutrient metal in order to colonize their hosts, and alterations in dietary metal levels profoundly affect susceptibility to infection. The host protein calprotectin (CP) is one of the most important contributors to immune-mediated metal restriction and CP protects against infection through the chelation of nutrient zinc (Zn) and manganese (Mn). In this application, we describe our use of CP as a probe to uncover a genetic locus within A. baumannii that is involved in survival during conditions of CP-dependent Zn starvation. This locus encodes a member of the conserved COG0523 family of GTPases that we have named Zur-induced GTPase A (ZigA), and a D-alanine D-alanine carboxypeptidase that we have named Zn-regulated lipoprotein A (ZrlA). We have discovered that ZigA is a metallochaperone that provides Zn to client proteins and is required for the liberation of a biovavailable Zn pool during conditions of Zn starvation. This finding establishes ZigA as the first example of a Zn metallochaperone in nature. In addition, our model predicts that ZrlA is required to maintain cell wall architecture during Zn stress. Based on these fundamental discoveries, we hypothesize that upon Zn starvation, A. baumannii mobilizes Zn to critical Zn-requiring client proteins, while also activating the expression of a carboxypeptidase involved in peptidoglycan remodeling that substitutes for the loss of activity of a Zn-requiring paralog. To test this central hypothesis, we propose a series of experiments aimed at understanding the mechanism and pathophysiological consequence of the A. baumannii response to dietary and host-imposed Zn deprivation during the pathogenesis of pneumonia. In these studies, we will (i) define the target of the ZigA Zn-metallochaperone during conditions of CP- imposed Zn restriction, (ii) elucidate the contribution of ZrlA to peptidoglycan remodeling during conditions of Zn deprivation, and (iii) determine the importance of Zn distribution during the pathogenesis of A. baumannii pneumonia. These results will provide fundamental insight into how A. baumannii responds to nutrient deprivation in the vertebrate host, and lay the foundation for the creation of peptide therapeutics based on a CP scaffold that inhibit microbial growth through nutrient metal chelation.