Mycobacterium turberculosis metal transport P-type ATPases

描述（由申请人提供）： 结核病是全球主要的健康问题。结核分枝杆菌的毒力是基于其驻留在巨噬细胞内的能力。宿主防御包括活性氧、活性氮、去除金属微量营养素和降低pH。微生物部分通过分泌各种金属酶（SOD、过氧化氢酶等）存活。来克服巨噬细胞的防御其致病策略的中心是处理所涉及的重金属（Cu+、Zn 2+、Fe 2+、Mn 2+等），质子和可能的碱金属如K+和Mg 2+。M.结核病基因组揭示了许多金属转运蛋白的存在。其中，P型ATP酶最为突出。M.结核病有12个编码这些蛋白质（命名为Ctp，阳离子转运蛋白）的基因：3个Cu+-ATP酶，1个K+-ATP酶，4个未知底物的重金属ATP酶和4个可能转运H+或碱金属的ATP酶。我们假设这些转运蛋白的病理生理作用主要由其底物特异性决定。这些P-型ATP酶的亚组的金属特异性将通过在E.大肠杆菌，检查细菌功能的获得，并测量酶和运输活性。先前的研究表明，CtpC和CtpD与体内（巨噬细胞）M特别相关。结核病增长。测试这个，M。结核突变体将与过表达这些蛋白的菌株一起沿着构建。将评估它们的毒力（在小鼠和培养的巨噬细胞中）和在各种培养基中生长的适应性。突变和过表达这些蛋白质对M.将确定结核金属稳态及其响应宿主氧化还原攻击的能力。这些研究的结果将提供这些蛋白质的重要性的初步评价，并使这些转运蛋白的生物学功能的产生有意义的可验证的假设。公共卫生相关性：世界上大约三分之一的人口感染结核分枝杆菌。尽管有可用的疫苗和有效的治疗方法，但每年仍有200万人死于结核病。该项目将描述一组对结核病毒力至关重要的蛋白质。

DESCRIPTION (provided by applicant): Tuberculosis is a leading health problem worldwide. Mycobacterium tuberculosis virulence is based on its capability to reside inside macrophages. Host defense includes reactive oxygen species, reactive nitrogen species, removal of metal micronutrients and lowered pH. The microorganism survives, in part, by secreting various metalloenzymes (SODs, catalases, etc.) to overcome macrophage defenses. At the center of its pathogenic strategy is the handling of involved heavy metals (Cu+, Zn2+, Fe2+, Mn2+, etc.), protons and perhaps alkali metals such as K+ and Mg2+. M. tuberculosis genome reveals the presence of numerous metal transporters. Among these, the more prominent are P-type ATPases. M. tuberculosis has 12 genes coding for these proteins (named Ctp, cation transport protein): 3 Cu+-ATPases, a K+-ATPase, 4 heavy metal ATPases of unknown substrate and 4 ATPases that likely transport H+ or alkali metals. We hypothesize that the pathophysiological role of these transporters is primarily determined by their substrate specificity. Metal specificities of a subset of these P-type ATPases will be determined by expressing these in E. coli, examining bacterial gain of function, and measuring the enzymatic and transport activity. Previous studies suggest that CtpC and CtpD are particularly relevant for in vivo (macrophage) M. tuberculosis growth. Testing this, M. tuberculosis mutants will be constructed along with strains overexpressing these proteins. Their virulence (in mice and cultured macrophages) and fitness to growth in various media will be assessed. The impact of mutating and overexpressing these proteins on M. tuberculosis metal homeostasis and its ability to respond to the host redox attack will be determined. Results form these studies will provide an initial appreciation of these proteins importance and enable the generation meaningful testable hypothesis on the biological function of these transporters. PUBLIC HEALTH RELEVANCE: Approximately one-third of the world's population is infected with Mycobacterium tuberculosis. In spite of an available vaccine and effective treatments, there are 2 million tuberculosis-related deaths per year. This project will characterize a group of proteins that appear essential for tuberculosis virulence.