Structural And Biochemical Characterization Of A Novel Mycobacterial Heme Uptake

新型分枝杆菌血红素摄取的结构和生化特征

• 批准号: 7867903
• 负责人: Celia Goulding
• 金额: $ 49.4万
• 依托单位: UNIVERSITY OF CALIFORNIA-IRVINE
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-06-15 至 2014-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7867903
• 关键词: Accounting; Affect; Affinity; Amino Acid Substitution; Antitubercular Agents; Attenuated; Bacteria; Binding; Biochemical; Carrier Proteins; Collaborations; Communities; Data; Development; Disease; Environment; Genes; Genus Mycobacterium; Goals; Heme; Heme Iron; Hemoglobin; Human; Investigation; Iron; Laboratories; Life; Light; Mass Spectrum Analysis; Membrane; Metals; Mind; Molecular; Mutation; Mycobacterium tuberculosis; Nuclear Magnetic Resonance; Pathway interactions; Play; Protein Sequence Homologs; Proteins; Proteome; Proteomics; Research; Resolution; Role; Siderophores; Source; Structure; System; Techniques; Transferrin; Tuberculosis; Tuberculosis Vaccines; base; drug development; extracellular; in vivo; killings; mutant; mycobacterial; mycobactins; novel; novel therapeutics; pathogen; prevent; public health relevance; therapeutic development; tool; tuberculosis drugs; uptake; yeast two hybrid system

DESCRIPTION (provided by applicant): Tuberculosis (TB) continues to kill millions of people around the world. New tools to prevent and treat this disease are urgently needed. Iron is an essential metal for all forms of life and most bacterial pathogens including mycobacteria must import iron from its host to survive. Hence iron acquisition pathways are well studied in mycobacteria as their components are essential to mycobacterial viability. Thus far, it is thought that iron uptake in mycobacteria is orchestrated by mycobactins that are capable of removing iron from human transferrin. However in humans, transferrin iron accounts for less than 1% of the body's total iron whereas heme iron can represent greater than 80%. Thus one may speculate that mycobacteria are capable of acquiring iron from human heme sources. Recent studies, which focus on mycobactin deficient mutants of Mycobacterium tuberculosis (Mtb) and BCG, suggest that there is a novel heme acquisition pathway in mycobacteria. Interestingly, BCG has an attenuated heme uptake pathway compared to Mtb. In addition, an Mtb proteome-wide approach has been undertaken in our laboratory to identify potential proteins involved in heme acquisition. We propose a putative pathway where heme is sequestered from human hemoglobin by a secreted hemophore, transferred across the membrane by heme transporters, and broken-down by cytosolic heme-degrading protein to release iron. This research will shed light on the molecular mechanism of heme transfer from humans to bacteria. In addition, we will investigate the affect each gene within this proposed pathway, has on mycobacterial heme uptake in vivo. The specific aims of this proposal are as follows: 1) Biophysical and biochemical investigation of the novel mycobacterial hemophore. 2) Explore the mechanism of heme transfer from host hemoglobin to hemophore to heme transporter. 3) Identification and characterization of other proteins involved in heme uptake. 4) Investigation into mycobacterial heme uptake system in Mtb. My proposed research focuses on gaining a comprehensive understanding of this novel mycobacterial heme uptake system on both molecular (single protein) and cellular levels. Interestingly, most of the proteins involved in this pathway have no close non-mycobacterial, protein sequence homologs, and additionally, the hemophore has a novel three-dimensional fold. With this in mind, the heme uptake pathway provides a number of good protein targets for the development of therapeutics against TB. PUBLIC HEALTH RELEVANCE: Tuberculosis (TB) continues to kill millions of people worldwide and is caused by the bacterial pathogen Mycobacterium tuberculosis (Mtb). Iron is essential for all living species, and the TB community thought that Mtb acquired its iron in vivo solely via its iron siderophores; thus our identification of a novel heme acquisition pathway whereby iron is obtained via the uptake of human heme breaks this paradigm. In this proposal, we will characterize this mycobacterial specific heme uptake pathway at a molecular and cellular level, which will open new avenues for anti-TB drug development.

描述（由申请人提供）：结核病（TB）继续在世界各地杀死数百万人。目前迫切需要预防和治疗这种疾病的新工具。铁是所有生命形式必不可少的金属，包括分枝杆菌在内的大多数细菌病原体必须从宿主那里进口铁才能生存。因此，铁获取途径在分枝杆菌中得到了很好的研究，因为它们的成分对分枝杆菌的生存能力至关重要。到目前为止，人们认为分枝杆菌中的铁摄取是由能够从人转铁蛋白中去除铁的分枝杆菌素精心安排的。然而，在人体中，转铁蛋白铁只占体内总铁的不到1%，而血红素铁却占80%以上。因此，人们可以推测分枝杆菌能够从人类血红素来源获取铁。最近对结核分枝杆菌（Mtb）和卡介苗的mycoactin缺陷突变体的研究表明，分枝杆菌中存在一种新的血红素获取途径。有趣的是，与结核分枝杆菌相比，卡介苗具有较弱的血红素摄取途径。此外，我们的实验室已经开展了一项Mtb蛋白质组范围的方法，以确定参与血红素获取的潜在蛋白质。我们提出了一种假设的途径，即血红素通过分泌的血红细胞从人血红蛋白中分离出来，通过血红素转运蛋白在细胞膜上转移，并被胞质血红素降解蛋白分解释放铁。这项研究将揭示血红素从人转移到细菌的分子机制。此外，我们将研究该途径中每个基因对分枝杆菌体内血红素摄取的影响。本课题的具体目的如下：1)新型分枝杆菌血球的生物物理生化研究。2)探讨血红素从宿主血红蛋白到红细胞再到血红素转运体的转运机制。3)与血红素摄取有关的其他蛋白质的鉴定和表征。结核分枝杆菌血红素摄取系统的研究。我提出的研究重点是在分子（单蛋白）和细胞水平上全面了解这种新型分枝杆菌血红素摄取系统。有趣的是，参与该途径的大多数蛋白质没有密切的非分枝杆菌蛋白序列同源物，此外，血团具有新颖的三维折叠。考虑到这一点，血红素摄取途径为开发治疗结核病的药物提供了许多良好的蛋白质靶点。公共卫生相关性：结核病（TB）继续导致全世界数百万人死亡，由细菌病原体结核分枝杆菌（Mtb）引起。铁对所有生物物种都是必需的，结核病界认为结核分枝杆菌仅通过铁铁载体在体内获得铁；因此，我们鉴定了一种新的血红素获取途径，通过摄取人类血红素获得铁，打破了这种范式。在本文中，我们将在分子和细胞水平上表征这种分枝杆菌特异性血红素摄取途径，这将为抗结核药物的开发开辟新的途径。