Bacterial Heme Transport by non-Isd NEAT Proteins

描述（由申请方提供）：为了在哺乳动物宿主中复制，细菌病原体必须获得宿主铁。然而，大多数铁被螯合在铁卟啉血红素中，铁卟啉血红素与循环血红蛋白（Hb）结合。为了进入这个铁池并增强血液中的复制，假设细菌需要专门的基于蛋白质的系统来释放和运输血红素穿过细胞包膜。 最近的研究表明，血红素收购革兰氏阳性菌是由一个结构模块称为近铁转运（NEAT）域介导的。NEAT蛋白是铁调节的表面决定簇（Isd）系统的标志，所述铁调节的表面决定簇（Isd）系统是起输入和分解代谢血红素的作用的转运蛋白的网络。然而，对测序的革兰氏阳性致病菌的基因组的分析表明，编码NEAT蛋白的基因也可以与Isd操纵子不同。这就提出了关于这些额外的NEAT蛋白在血红素摄取过程中的作用的问题。 本申请试图确定两种非Isd NEAT蛋白BslK和BAS 0520在B中的功能。炭疽病发病机制和血红素输入。此外，这些NEAT从宿主血红素库获得血红素并启动血红素转移到细胞中的分子机制也将被确定。我们假设BslK和BAS 0520促进血红素从血红蛋白（Hb）提取并将血红素转移到包膜蛋白，从而增强B。炭疽菌在宿主感染期间复制。将以下列具体目标检验这一假设： 1.确定非ISD NEAT蛋白对炭疽发病机制和血红素输入的贡献。 - B。将测试缺乏一种或两种基因的炭疽菌株在作为唯一铁源的Hb或血红素上的生长，并使用炭疽病的完全毒性模型确定它们在感染期间的作用。 2.确定BslK和0520介导的血红素获得的机制，并确定参与血红素同化的新基因。 - 新的生物物理学方法将用于确定BslK和BAS 0520功能的机制。转座子突变库将被筛选，以发现新的血红素转运系统。 公共卫生相关性：这项研究调查了一类铁吸收蛋白的作用机制，这些蛋白有助于某些细菌物种的生存和致病能力。从这些研究中产生的知识将增加我们对细菌病原体铁获取的理解，促进针对这些系统的新型抗感染药物的开发，并产生用于创建更安全有效疫苗的试剂。

DESCRIPTION (provided by applicant): To replicate in mammalian hosts, bacterial pathogens must acquire host iron. Most iron, however, is sequestered in the iron-porphyrin heme which is bound to circulating hemoglobin (Hb). To access this iron pool and enhance replication in blood, it is hypothesized bacteria require specialized protein-based systems to liberate and transport the heme across the cell envelope. Recent studies have demonstrated that heme acquisition in Gram-positive bacteria is mediated by a structural module called the near-iron transporter (NEAT) domain. NEAT proteins are a hallmark of iron- regulated surface determinant (Isd) systems, a network of transport proteins that function to import and catabolize heme. However, analysis of the genomes of sequenced Gram-positive pathogenic bacteria indicates genes encoding NEAT proteins can also be distinct from Isd operons. This raises the question as to the role of these additional NEAT proteins in the heme uptake process. This application seeks to determine the function of two non-Isd NEAT proteins, BslK and BAS0520, in B. anthracis pathogenesis and heme import. Further, the molecular mechanisms by which these NEATs acquire heme from host heme reservoirs and initiate heme transfer into the cell will also be determined. We hypothesize BslK and BAS0520 promote heme extraction from hemoglobin (Hb) and transfer the heme to envelope proteins, thereby enhancing B. anthracis replication during host infection. This hypothesis will be tested with the following specific aims: 1. Determine the contribution of non-Isd NEAT proteins to anthrax pathogenesis and heme import. --- B. anthracis strains lacking one or both genes will be tested for growth on Hb or heme as the sole iron source and their role during infection determined using fully-virulent models of anthrax disease. 2. Determine the mechanism of BslK- and 0520-mediated heme acquisition and identify new genes involved in heme assimilation. --- Novel biophysical methods will be used to determine the mechanism of BslK and BAS0520 function. A transposon mutant library will be screened to uncover new heme transport systems. PUBLIC HEALTH RELEVANCE: This study investigates the mechanism of action of a class of iron-uptake proteins that contribute to the survival and disease-causing ability of certain bacterial species. Knowledge generated from these studies will increase our understanding of iron acquisition in bacterial pathogens, facilitate the development of novel anti-infectives targeting these systems, and generate reagents for the creation of more safe and effective vaccines.