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: 8367291
• 负责人: Erica Francesca Miller
• 金额: $ 3.09万
• 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 2013-08-22
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8367291
• 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.

描述(申请人提供)：幽门螺杆菌由S在20世纪80年代初发现，是消化性和十二指肠溃疡、胃炎、贫血和某些胃癌的常见病因。这种细菌通常是在儿童时期获得的，拥挤的生活条件和经济差距与较高的感染率密切相关。尿素酶是幽门螺杆菌感染所必需的一种酶，占细胞总蛋白的近10%。这种酶必须是活性的，才能使细菌在宿主上定居，这一事实已被尿素酶中和和缓冲胃酸的能力所解释。然而，在定植中对尿素酶的需求不取决于酸性条件，这表明该酶的替代作用不只是简单的耐酸。这项研究计划的主要目标是研究尿素酶的这些替代作用之一-它在氮代谢中的作用。在过去的15年里，尿素酶研究中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，即尿素酶是一种细胞内酶，与以前的看法相反。其次，通过对15N-尿素的示踪研究，证明了尿素氮已进入细胞内。此外，高浓度的尿素还被证明可以杀死幽门螺杆菌，这种作用是由尿素的水解和随后细胞内铵的积累来解释的。尿素氮的同化机制很少通过生化方法来研究，然而，初步的基因组水平的比较表明，幽门螺杆菌的氨同化途径并不是其他蛋白细菌的典型途径。事实上，幽门螺杆菌在所有其他研究得很好的细菌分类群中是一个反常的物种，无论是在结构上还是在调节这种有机体中被认为同化铵的酶-谷氨酰胺合成酶和谷氨酸脱氢酶方面。此外，谷氨酰胺合成酶可能是与尿素酶相互作用的伙伴，暗示了尿素酶和氨同化之间潜在的密切关系。本研究利用诱变等分子技术，结合酶纯化和动力学研究等生化技术，研究尿素从胃粘膜直接进入幽门螺杆菌初级代谢中心的途径。