Mechanistic Investigations of Helicobacter pylori Chemotaxis

幽门螺杆菌趋化性的机制研究

• 批准号: 8081779
• 负责人: Emily Sarah Marie Sweeney
• 金额: $ 5.13万
• 依托单位: UNIVERSITY OF OREGON
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-07-01 至 2013-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8081779
• 关键词: Acids; Affect; Ammonia; Antibiotics; Bacteria; Behavior; Behavioral Assay; Binding; Binding Sites; Biochemical; Biological Assay; Cancer Etiology; Carbon Dioxide; Carcinogens; Cause of Death; Cessation of life; Chemicals; Chemoreceptors; Chemotaxis; Complex; Crystallography; Cues; Detection; Drug Design; Environment; Epithelial; Epithelium; Escherichia coli; Helicobacter Infections; Helicobacter pylori; In Vitro; Incidence; Infection; Investigation; Knowledge; Laboratories; Lead; Malignant Neoplasms; Membrane; Molecular Conformation; Mutation; Periplasmic Binding Proteins; Pharmaceutical Preparations; Population; Prevalence; Protein Binding; Research Design; Resolution; Risk Factors; Role; Rotation; Signal Transduction; Stomach; Structure; Swimming; Testing; Urea; VAI-2; Vibrio; Work; base; design; expression vector; in vivo; interest; malignant stomach neoplasm; mutant; novel; periplasm; prevent; protonation; quorum sensing; response

DESCRIPTION (provided by applicant): Helicobacter pylori inhabit the highly acidic stomach and infection can lead to gastric cancer. In order for H. pylori to survive in this environment, it must detect chemical cues such as acid. These cues direct H. pylori to the stomach epithelium. We and others have shown that the chemoreceptor TlpB is responsible for detecting HCl. We have determined the atomic structure of the periplasmic domain of TlpB and found, to our surprise, that urea is bound. H. pylori uses urea to neutralize its environment and survive in stomach acid. We have found that urea stabilizes TlpB and may be part of the mechanism by which H. pylori senses acid. We have also shown that TlpB detects the bacterial quorum sensing molecule autoinducer 2 (AI-2), that we suspects promotes H. pylori dispersal throughout the stomach. TlpB does not interact directly with AI-2, however, we have identified five candidate periplasmic binding proteins that may. A similar AI-2 detection mechanism has been shown in Vibrio Harveyi where a periplasmic protein binds AI-2 and then interacts with a membrane bound chemoreceptor, similar to TlpB. Determining the mechanisms of H. pylori chemotaxis to acid and AI-2 will aid in designing novel antibiotics to treat H. pylori infection and decrease the incidence of gastric cancer. Objective/Hypothesis: Previous studies indicate that the chemoreceptor TlpB is responsible for H. pylori chemotaxes away from acid and AI-2. We propose to determine the mechanisms by which TlpB detects and responds to both acid and AI-2. Specific Aims: (1) We will test the hypothesis that TlpB responds to acid by binding urea in a pH sensitive manner; (2) we will test the hypothesis that TlpB responds to AI-2 through interactions with a periplasmic binding protein that binds AI-2. Study Design: First, we will determine if urea binds and stabilizes TlpB in a pH sensitive manner. We will then identify mutations in TlpB that disrupt urea binding and prevent proper acid chemotaxis. These TlpB mutants will be used in crystallography studies to identify conformation changes in the presence or absence of urea. To determine TlpB's role in sensing AI-2, we have identified candidate periplasmic binding proteins that may bind AI-2 and, when deleted in H. pylori, may be defective in AI-2 chemotaxis. We will then determine whether the candidate proteins bind AI-2 and/or TlpB directly. The structures of the periplasmic binding proteins of interest with AI-2 and TlpB will be pursued. Cancer Relevance: These studies will contribute to our understanding of how H. pylori senses chemical cues within the environment of the stomach. The detection of these cues by H. pylori is vital for its ability to infect the stomach and promote gastric cancer. More knowledge of the mechanisms of acid and AI-2 chemotaxis will provide clear targets for drug design and ultimately decrease the prevalence of gastric cancer.

描述(申请人提供)：幽门螺杆菌栖息在高酸性的胃部，感染可导致胃癌。为了让幽门螺杆菌在这种环境中生存，它必须检测到酸等化学线索。这些线索将幽门螺杆菌导向胃上皮细胞。我们和其他人已经证明，化学感受器TlpB负责检测HCl。我们已经确定了TlpB的周质结构域的原子结构，并发现令我们惊讶的是，尿素是结合的。幽门螺杆菌使用尿素中和环境，并在胃酸中存活。我们发现尿素稳定了TlpB，这可能是幽门螺杆菌感酸机制的一部分。我们还证明了TlpB检测到细菌群体感应分子自动诱导因子2(AI-2)，我们怀疑AI-2促进了幽门螺杆菌在胃中的扩散。TlpB不直接与AI-2相互作用，然而，我们已经确定了五个可能与AI-2相互作用的候选周质结合蛋白。在哈维氏弧菌中也显示了类似的AI-2检测机制，其中周质蛋白与AI-2结合，然后与膜结合的化学受体相互作用，类似于TlpB。确定幽门螺杆菌对酸和AI-2的趋化机制将有助于设计新的抗生素来治疗幽门螺杆菌感染，降低胃癌的发病率。目的/假设：以前的研究表明，化学受体TlpB负责幽门螺杆菌的趋化作用，远离酸性和AI-2。我们建议确定TlpB检测和响应酸和AI-2的机制。具体目标：(1)我们将测试TlpB通过以pH敏感的方式结合尿素来响应酸的假设；(2)我们将测试TlpB通过与结合AI-2的周质结合蛋白的相互作用来响应AI-2的假设。研究设计：首先，我们将确定尿素是否以pH敏感的方式结合和稳定TlpB。然后，我们将确定TlpB中破坏尿素结合并阻止适当的酸性趋化的突变。这些TlpB突变体将用于结晶学研究，以确定尿素存在或不存在时的构象变化。为了确定TlpB在感知AI-2中的作用，我们已经确定了候选的周质结合蛋白，这些蛋白可能与AI-2结合，当在幽门螺杆菌中缺失时，可能在AI-2的趋化作用中存在缺陷。然后我们将确定候选蛋白是否直接与AI-2和/或TlpB结合。我们将继续研究与AI-2和TlpB相关的周质结合蛋白的结构。癌症相关性：这些研究将有助于我们了解幽门螺杆菌如何感知胃环境中的化学线索。幽门螺杆菌对这些信号的检测对于它感染胃和促进胃癌的能力至关重要。更多地了解ACID和AI-2的趋化机制将为药物设计提供明确的靶点，并最终降低胃癌的发病率。