Heme utilization and homeostasis in Pseudomonas aeruginosa

描述（由申请方提供）：病原菌需要铁才能存活并引起感染。许多细菌病原体已经进化出复杂的系统来利用血红素作为铁的主要来源。机会致病菌铜绿假单胞菌是医院感染的一个越来越常见的原因，与这种感染相关的发病率很高。除了它们对许多抗生素的天然耐药性外，此类感染的惊人增加突出了替代抗菌策略的必要性。在以前的生化 和生物物理学研究，我们已经表征了细胞内血红素运输蛋白H2S和铁调节的血红素加氧酶HemO，并进一步表明它们对血红素利用和毒性至关重要。该提案的具体目标是进一步阐明细胞外血红素利用和调节的机制，因为它与毒力有关。这将是 通过实现; i）通过遗传、生物化学和代谢组学方法阐明细胞内血红素运输途径并定义胆绿素IX-δ和β异构体在血红素摄取的细胞外反馈调节中的作用。ii）通过定点诱变、生物化学和生物物理学研究的组合来定义血红素从Hem S转移到HemO的机制，以及iii）确定调节性小RNA PrrH在血红素摄取和利用中的作用。这些研究将为细菌病原体中血红素和铁的稳态提供新的见解。我们将通过这一建议，确定新的血红素代谢和调节途径作为新的抗菌目标，并推进目前的理解血红素转移机制。

DESCRIPTION (provided by applicant): Pathogenic bacteria require iron for their survival and ability to cause infection. Many bacterial pathogens have evolved sophisticated systems to utilize heme as a primary source of iron. The opportunistic pathogen P. aeruginosa is an increasingly common cause of nosocomial infections with high morbidity associated with such infections. In addition to their natural resistance to many antibiotics the alarming increase in such infections highlights the need for alternate antimicrobial strategies. In previous biochemical and biophysical studies we have characterized the intracellular heme trafficking protein PhuS and the iron-regulated heme oxygenase HemO and have further shown they are critical for heme utilization and virulence. The specific aims of the proposal are to further elucidate the mechanism of extracellular heme utilization and regulation as it relates to virulence. This will be achieved through; i) Elucidating the intracellular heme trafficking pathway and defining the role of biliverdin IX-delta and -beta isomers in the extra-cellular feedback regulation of heme uptake through genetic, biochemical, and metabolomics approaches. ii) Defining the mechanism of heme transfer from PhuS to HemO through a combination of site-directed mutagenesis, biochemical and biophysical studies and iii) Determining the role of the regulatory small RNA PrrH in heme uptake and utilization. These studies will provide new insights into heme and iron homeostasis in bacterial pathogens. We will through this proposal identify new heme metabolic and regulatory pathways as novel antimicrobial targets and advance the current understanding of heme transfer mechanisms.