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）：在1980年代初期发现的幽门螺杆菌是消毒和十二指肠溃疡，胃炎，贫血和某些胃癌的常见原因。该细菌通常是在儿童时期获得的，具有拥挤的生活条件和经济差异与较高的感染率密切相关。脲酶是幽门螺杆菌感染所需的一种酶，占总细胞蛋白的近10％。该酶必须活跃，以使细菌定居其宿主，这一事实是由尿素释放的能力中和和缓冲液抵抗胃酸性的。然而，在定植中尿素酶的要求不包括酸性条件，这表明该酶除了简单的抗酸性外，其他作用。 该研究计划的主要目的是检查这些替代作用之一，用于脲酶在氮代谢中的作用。在过去的十五年中，尿素研究中的两个关键转折点确立了这种酶为氮代谢的球。首先，一组发现导致当前的范例是尿布是一种细胞内酶，与以前持有的信念相反。其次，通过追踪15N-rea的研究，尿素氮被证明被掺入细胞中。此外，高浓度的尿素被证明会杀死幽门螺杆菌，这种作用是通过尿素的水解以及随后在细胞中铵的积累来解释的。 几乎无法通过生化方法检查尿素氮的吸收机制。然而，初步的基因组水平比较表明，幽门螺杆菌中铵同化的途径并不是其他蛋白质细菌的典型特征。实际上，幽门螺杆菌在所有其他良好的细菌分类群中都是一种异常，就其结构和调节而言，这些酶在这种有机体 - 谷氨酰胺合酶和谷氨酸脱氢酶中被认为是铵的调节。此外，谷氨酰胺合成酶是尿素酶的推定相互作用伴侣，暗示了尿素酶和铵同化之间的潜在紧密关系。本研究使用分子技术（例如诱变技术）以及诸如酶纯化和动力学研究之类的生化技术，旨在将将尿素直接从胃mileu直接带到幽门螺杆菌中心的心脏的途径来表征。 公共卫生相关性：在美国的某些地区，将近一半的人口长期感染了幽门螺杆菌，幽门螺杆菌是I类致癌物和胃溃疡的病因。尿布酶是幽门螺杆菌定殖所必需的，但该酶仍未在细菌生理和代谢的晶状体下进行研究。为了了解该细菌与宿主的相互作用如何取决于对尿素酶及其在发病机理中的作用的完全理解。