Molecular mechanisms of antibiotic resistance

抗生素耐药性的分子机制

• 批准号: 7064635
• 负责人: FOCCO VAN DEN AKKER
• 金额: $ 27.04万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-07-01 至 2007-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7064635
• 关键词: Raman spectrometry; X ray crystallography; active sites; antibiotics; bacteria; bacterial genetics; bacterial proteins; beta lactam antibiotic; beta lactamase; drug resistance; enzyme inhibitors; enzyme mechanism; gene mutation; hydrolysis; protein structure; structural biology

Bacterial infections pose serious threats to human health. Furthermore, viral infections such as the flu are frequently accompanied by bacterial infections which is often a deadly combination. Due to the development of resistance, the options for treating infections have dwindled substantially. This resistance is in large part due to the bacterial expression of beta-lactamases that degrade beta-lactam antibiotics. Our main goal is to structurally understand the molecular basis of resistance against the clinically available ceftazidime antibiotic and how certain mutations in beta-lactamases confer resistance to this antibiotic and other mutations confer resistance to the beta-lactamase inhibitors tazobactam, sulbactam, and clavulanic acid. Our structure-function studies involve a novel synergy between X-ray and time-resolved Raman crystallography. Our innovative inter-disciplinary approach allows us to identify and track reaction intermediates inside crystals prior to X-ray analysis and provides a unique advantage to accomplish our Specific Aims: Specific Aim 1: To test the hypothesis that SHV-1 resistance variants at position M69 cause active site changes that affect the level and stability of the intermediate formation involving the 3 inhibitors thus allowing them to overcome the inhibition. Specific Aim 2: To test the hypothesis that the extended-spectrum beta-lactamase SHV-2 variant (G238S) and SHV-5 variant (G238S & E240K) have evolved to hydrolyze the third-generation cephalosporin ceftazidime by widening the active site and forming novel interactions with this compound. Specific Aim 3: To test the hypothesis that the changes at position D179 which are present in SHV-6, -8, and -24 have evolved to hydrolyze ceftazidime by shifting the omega loop thereby extending the active site to accommodate ceftazidime. The structural knowledge gained from the proposed research could aid in the design of new antibiotic drugs.

细菌感染对人类健康构成了严重威胁。 此外，流感等病毒感染经常伴有细菌感染，这通常是致命的组合。由于抗药性的发展，治疗感染的选择大大减少了。这种抗性在很大程度上是由于β-内酰胺酶的细菌表达降解β-内酰胺抗生素。我们的主要目标是在结构上了解对临床可用头孢菌素抗生素的抗性的分子基础，以及β-内酰胺酶中的某些突变如何赋予该抗生素的耐药性和其他突变赋予β-内酰胺酶抑制剂Tazobactam，sulbactam和clavulanic acter抗性。我们的结构功能研究涉及X射线和时间分辨的拉曼晶体学之间的新型协同作用。我们创新的跨学科方法使我们能够在X射线分析之前识别和跟踪晶体内部的反应中间体，并为实现我们的特定目的提供了独特的优势：特定目标1：测试以下假设：SHV-1抗性在位置M69处的抗性变体引起主动部位变化，影响涉及3种抑制剂的中间形成的水平和稳定性，从而使它们能够克服抑制作用。具体目的2：检验以下假设：扩展的频谱β-内酰胺酶SHV-2变体（G238S）和SHV-5变体（G238S＆E240K）已进化以通过拓宽活跃的现场和形成活跃的现场和形成活性的位点和形成，以水解第三代CEPHALOSPORIN CETFEFTAZIDIME与这种化合物的新颖相互作用。具体目的3：测试以下假设：SHV -6，-8和-24中存在的位置D179的变化已通过移动omega循环的变化而演变为水解头孢心夏，从而扩展了活跃的位点以容纳头孢心兹维时。从拟议的研究中获得的结构知识可以帮助设计新的抗生素药物。