Urease Activity and Nitrogen Assimilation in Helicobacter pylori

幽门螺杆菌的脲酶活性和氮同化

• 批准号: 8062828
• 负责人: Erica Francesca Miller
• 金额: $ 3.05万
• 依托单位: UNIVERSITY OF GEORGIA
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-08-23 至 2014-08-22
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8062828
• 关键词: Acclimatization; Accounting; Acidity; Acids; Address; Amino Acids; Ammonia; Ammonium; Anemia; Assimilations; Bacillus subtilis; Bacteria; Bacterial Physiology; Belief; Biochemical; Buffers; Carbon Dioxide; Carcinogens; Cells; Childhood; Crowding; Duodenal Ulcer; Economic Conditions; Enterobacter aerogenes bacterium; Environment; Enzymes; Epithelial Cells; Gastric ulcer; Gastritis; Genome; Genomics; Glutamate Dehydrogenase; Glutamate-Ammonia Ligase; Heart; Helicobacter Infections; Helicobacter pylori; Homologous Gene; Hydrolysis; In Vitro; Infection; Kinetics; Life; Metabolism; Methods; Molecular; Mutagenesis; Nitrogen; Nutrient; Organism; Outcome; Pathogenesis; Pathway interactions; Peptic Ulcer; Population; Property; Proteins; Proteobacteria; Protons; Regulation; Research; Research Training; Resistance; Role; Stomach; Structure; Surface; System; Taxon; Techniques; United States; Urea; Urea Nitrogen; Urease; cell motility; in vivo; killings; lens; malignant stomach neoplasm; nitrogen metabolism; novel; periplasm

DESCRIPTION (provided by applicant): Discovered in the early 1980's, Helicobacter pylori is a common cause of peptic and duodenal ulcers, gastritis, anemia, and certain gastric cancers. This bacterium is typically acquired during childhood, with crowded living conditions and economic disparities closely correlating with higher infection rates. Urease is an enzyme that is required for infection by H. pylori, and accounts for nearly 10% of the total cellular protein. This enzyme must be active in order for the bacterium to colonize its host, a fact that has been explained by urease's ability to neutralize and buffer against the acidity of the stomach. However, the requirement for urease in colonization is not contingent upon acidic conditions, indicating alternative roles for this enzyme other than simply acid resistance. The primary objective of this research plan is to examine one of these alternative roles for urease-its role in nitrogen metabolism. Over the past fifteen years, two crucial turning points in urease research established this enzyme as player in nitrogen metabolism. First, a set of discoveries led to the current paradigm that urease is an intracellular enzyme, contrary to previously held beliefs. Secondly, urea nitrogen was shown to be incorporated into the cell through tracing studies of 15N-urea. High concentrations of urea have furthermore been shown to kill H. pylori, an effect that was explained by the hydrolysis of urea and subsequent accumulation of ammonium in the cell. The mechanism by which urea nitrogen is assimilated has been scarcely examined through biochemical methods; however, preliminary genome-level comparisons revealed that the pathway of ammonium assimilation in H. pylori is not typical of other proteobacteria. In fact, H. pylori is an anomaly among all other well-studied bacterial taxa with regard to both the structure and the regulation of the enzymes thought to assimilate ammonium in this organism-glutamine synthetase and glutamate dehydrogenase. Furthermore, glutamine synthetase is a putative interacting partner with urease, alluding to a potentially tight relationship between urease and ammonium assimilation. Using molecular techniques such as mutagenesis, in conjunction with biochemical techniques such as enzyme purification and kinetic studies, this study aims to characterize the pathway that takes urea from the gastric mileu directly to the heart of H. pylori primary metabolism. PUBLIC HEALTH RELEVANCE: In some regions of the United States, nearly half of the population is chronically infected with Helicobacter pylori, a Class I carcinogen and causative agent of gastric ulcers. Urease enzyme is required for colonization of H. pylori, yet this enzyme still has not been studied under the lens of bacterial physiology and metabolism. To understand how this bacterium interacts with its host is contingent upon a complete understanding of urease and its role in pathogenesis.

描述（由申请人提供）：幽门螺杆菌发现于20世纪80年代早期，是消化性溃疡和十二指肠溃疡、胃炎、贫血和某些胃癌的常见原因。这种细菌通常在儿童时期获得，拥挤的生活条件和经济差距与较高的感染率密切相关。尿素酶是H. pylori，占总细胞蛋白的近10%。这种酶必须是活性的，以便细菌在其宿主中定居，这一事实已经被脲酶的中和和缓冲胃酸的能力所解释。然而，在定殖中对脲酶的要求并不取决于酸性条件，这表明这种酶除了简单的耐酸性之外还有其他作用。 本研究计划的主要目的是研究尿素酶在氮代谢中的作用。在过去的15年里，脲酶研究的两个关键转折点确立了这种酶在氮代谢中的作用。首先，一系列发现导致了当前的范式，即脲酶是一种细胞内酶，与以前持有的信念相反。其次，通过对15 N-尿素的示踪研究表明，尿素氮被掺入细胞中。此外，高浓度的尿素已被证明能杀死H。幽门螺杆菌，这是一种效果，这是由尿素的水解和随后的铵在细胞中的积累解释。 尿素氮的同化机制还很少通过生物化学方法研究，然而，初步的基因组水平的比较表明，在H.幽门不是其他变形菌的典型特征。事实上，H.幽门螺杆菌在所有其他研究充分的细菌分类群中，就其结构和被认为在该生物体中同化铵的酶-谷氨酰胺合成酶和谷氨酸脱氢酶的调节而言是异常的。此外，谷氨酰胺合成酶是一个假定的相互作用伙伴与尿素酶，暗示尿素酶和铵同化之间的潜在密切关系。利用分子技术，如诱变，结合生化技术，如酶纯化和动力学研究，本研究的目的是表征尿素从胃环境直接到心脏的H。幽门初级代谢。 公共卫生关系：在美国的一些地区，近一半的人口是慢性感染幽门螺杆菌，一类致癌物质和胃溃疡的病原体。尿素酶是H. pylori，但这种酶仍然没有在细菌生理学和代谢的透镜下进行研究。要了解这种细菌如何与其宿主相互作用，取决于对尿素酶及其在发病机制中的作用的全面了解。