Nitrosative stress defenses in periodontopathogen Porphyromonas gingivalis

牙周病原菌牙龈卟啉单胞菌的亚硝化应激防御

• 批准号: 9057873
• 负责人: Janina P Lewis
• 金额: $ 38.13万
• 依托单位: VIRGINIA COMMONWEALTH UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-06-26 至 2018-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9057873
• 关键词: Adult; Anti-Bacterial Agents; Bacteria; Binding; Bioinformatics; Biological Process; Cells; Code; Complex; Crystallography; DNA; DNA Binding; DNA Sequence; Defense Mechanisms; Environment; Flavodoxin; Gene Targeting; Genes; Growth; Health; Homeostasis; Human; Hydroxylamine; Invaded; Knowledge; Light; Mediating; Microarray Analysis; Molecular; Molecular Genetics; Molecular Structure; Nitric Oxide; Nitrite Reductase; Nitrites; Oral; Oral cavity; Organism; Oxidoreductase; Periodontal Diseases; Play; Porphyromonas gingivalis; Process; Property; Proteins; Proteome; Proteomics; Regulation; Regulon; Role; Signal Transduction; Structure; Therapeutic Agents; Tissues; Work; base; design; gene product; microorganism; mutant; nitrogen metabolism; nitrosative stress; novel; pathogen; periodontopathogen; prevent; tool; transcriptome; translational approach; weapons

DESCRIPTION (provided by applicant): Nitrite and nitric oxide are widespread and abundant in humans and are emerging as a potential new antibacterial therapeutic agents. The oral cavity has particularly high concentrations of nitrite, which can reach 1mM. Oral microorganisms have adapted to survive such high nitrosative stress exposure and disruption of these adaptation mechanisms would be expected to reduce growth and survival of bacteria in the oral environment. Porphyromonas gingivalis, a periodontopathogen, is well known for its high tolerance of nitrosative stress. However, the molecular basis of this tolerance is under-investigated. Using bioinformatics approaches and microarray analysis of P. gingivalis exposed to nitrite and nitric oxide, we identified several potential candidates that may play a role in nitrosative stress protection. The major upregulated gene was hcp, which codes for a putative hydroxylamine reductase. We have further identified a regulator, designated HcpR, which mediates expression of hcp and is required for growth of P. gingivalis in the presence of both nitrite and nitric oxide. We hypothesize that HcpR is a major player in adaptation of P. gingivalis to nitrosative stress. To determine its role in nitrosative stress protection, we will first definethe regulon of HcpR and the minimum DNA sequence required for its binding. Furthermore, we will carry out molecular structure studies of HcpR and of its complexes with DNA using NMR and crystallography. Since our study shows that the major regulated gene in P. gingivalis is hcp, we will determine the role of its gene product in adaptation of the bacterium to nitrosative stress an define its biological function in such adaptation. Also, P. gingivalis codes for multiple players that may be involved in nitrosative stress protection or nitrogen metabolism. We will characterize and determine the roles of those players in adaptation of P. gingivalis to nitrosative stress. Finally, we will investigate the role of nitrosative stress in host-pathogen interactions. The results of our study are expected to provide information regarding nitrosative stress homeostasis mechanisms in P. gingivalis at the regulatory and structural levels This knowledge will provide the tools to design agents that compromise the defense mechanisms of the periodontopathogen and turn endogenous human host nitrite and nitric oxide into a weapon that inhibits growth and ultimately can be exploited to treat periodontal disease. We predict that this work will shed light on nitrosative stress homeostasis mechanisms in a variety of other bacteria that carry similar nitrosative stress protection mechanisms to those in P. gingivalis.

描述（由申请人提供）：亚硝酸盐和一氧化氮在人类中广泛且丰富，并且正在成为潜在的新型抗菌治疗剂。口腔中的亚硝酸盐浓度特别高，可达1mM。口腔微生物已经适应了如此高的亚硝化应激暴露，并且这些适应机制的破坏预计会减少口腔环境中细菌的生长和存活。牙龈卟啉单胞菌是一种牙周病原菌，以其对亚硝化应激的高耐受性而闻名。然而，这种耐受性的分子基础尚未得到充分研究。利用生物信息学方法和对暴露于亚硝酸盐和一氧化氮的牙龈卟啉单胞菌进行微阵列分析，我们确定了几种可能在亚硝化应激保护中发挥作用的潜在候选者。主要上调的基因是 hcp，它编码假定的羟胺还原酶。我们进一步鉴定了一种调节剂，命名为 HcpR，它介导 hcp 的表达，并且是牙龈卟啉单胞菌在亚硝酸盐和一氧化氮存在下生长所必需的。我们假设 HcpR 是牙龈卟啉单胞菌适应的主要参与者 亚硝化应激。为了确定其在亚硝化应激保护中的作用，我们首先定义 HcpR 的调节子及其结合所需的最小 DNA 序列。此外，我们将使用核磁共振和晶体学对 HcpR 及其与 DNA 的复合物进行分子结构研究。由于我们的研究表明牙龈卟啉单胞菌中的主要调控基因是hcp，因此我们将确定其基因产物在细菌适应亚硝化应激中的作用，并定义其在这种适应中的生物学功能。此外，牙龈卟啉单胞菌编码可能参与亚硝化应激保护或氮代谢的多个参与者。我们将描述并确定这些参与者在牙龈卟啉单胞菌适应亚硝化过程中的作用 压力。最后，我们将研究亚硝化应激在宿主-病原体相互作用中的作用。我们的研究结果预计将提供有关牙龈卟啉单胞菌在调节和结构水平上的亚硝化应激稳态机制的信息。这些知识将提供工具来设计破坏牙周病原体防御机制的药物，并将内源性人类宿主亚硝酸盐和一氧化氮转化为抑制生长的武器，最终可用于治疗牙周病。我们预测这项工作将揭示多种其他细菌的亚硝化应激稳态机制，这些细菌与牙龈卟啉单胞菌具有相似的亚硝化应激保护机制。