Conformational Flexibility and Antibiotic Resistance

构象灵活性和抗生素耐药性

• 批准号: 8116653
• 负责人: JEFFREY W PENG
• 金额: $ 22.05万
• 依托单位: UNIVERSITY OF NOTRE DAME
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-08-24 至 2013-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8116653
• 关键词: Acceleration; Accounting; Acylation; Affect; Affinity; Amino Acids; Anti-Infective Agents; Antibiotic Resistance; Antibiotics; Automobile Driving; Binding; Carbapenems; Cephalosporins; Clinic; Clinical; Containment; Data; Dissection; Drug resistance; Event; Exposure to; Failure; Goals; Investigation; Lactamase; Lactams; Length; Ligands; Light; Link; Mass Spectrum Analysis; Membrane; Methods; Molecular; Molecular Conformation; Monobactams; Multi-Drug Resistance; Multienzyme Complexes; Nosocomial Infections; Nuclear Magnetic Resonance; Pattern; Penicillins; Peptides; Phage Display; Principal Investigator; Production; Protein Dynamics; Proteins; Public Health; Resistance; Resolution; Roentgen Rays; Science; Scourge; Sensory; Series; Signal Transduction; Site; Solutions; Structure; Transducers; Work; base; combat; conformational conversion; coping; drug resistant bacteria; extracellular; flexibility; inhibitor/antagonist; methicillin resistant Staphylococcus aureus; pathogen; programs; protein structure; public health relevance; receptor; resistance mechanism; sensor; tool

DESCRIPTION (provided by applicant): Methicillin-resistant Staphylococcus aureus (MRSA) is a multi-drug resistant bacteria pathogen that has become a global clinical threat, accounting for nearly one-third of hospital-acquired infections. The MRSA threat is especially insidious due to its resistance to 2-lactam antibiotics (e.g. penicillins, carbapenems, cephalosporins), which remain the most widely used anti-infectives in the clinic. Elucidating the molecular basis for MRSA drug resistance is therefore imperative to develop strategies for its containment. A principal resistance mechanism of MRSA is the production of a ?-lactamase that hydrolytically destroys ?-lactam antibiotics. This production is triggered by the transmembrane sensor/transducer protein BlaR1. Exposure to a ?-lactam antibiotic triggers signal transduction by BlaR1, which leads to ?-lactamase production. Recent CD and IR studies demonstrate that the signal transduction entails conformational transitions in the extracellular sensor domain of BlaR1 (BlaRS henceforth) upon its binding of ?-lactam antibiotics. Defining these conformational transitions at the atomic level has become a critical goal in efforts to elucidate the signal transduction mechanism. Despite the availability of high-resolution protein structures, the atomic-level mechanism for the conformational transitions driving BlaR1 signal transduction remains obscure. We have begun studies of BlaRS, and our preliminary results point to a new set of molecular factors important for the BlaRS conformational transitions: protein conformational dynamics. Specifically, using solution Nuclear Magnetic Resonance (NMR), we observe that ?-lactam binding causes site-specific changes in local BlaRS flexibility. Hence, to elucidate the key events underlying BlaR1 signal transduction, we will investigate the functional consequences of BlaRS conformational dynamics. Accordingly, we propose three Specific Aims that will define how the interplay between protein dynamics and conformational change facilitates signal transduction on the part of BlaRS. These Aims include: (i) Comparing the site-specific changes in BlaRS flexibility upon activation by a series of different ?-lactam substrates; (ii) Defining the mechanism of interaction between BlaRS, and a peptide derived from the extra-cellular Loop 2 of the trans-membrane region of the BlaR1 receptor; (iii) Compare the effects of non-??-lactam inhibitors of BlaR1 versus ?-lactam substrates (antibiotics), on the dynamics and conformation of BlaRS. Our main experimental tool will be multi-dimensional NMR, which provides a uniquely powerful method for the atomic-level description of protein dynamics on a per- amino-acid-residue basis. Our studies will shed new light on the inner-workings of the signal transduction mechanism responsible for pernicious ?-lactam antibiotic resistance in MRSA. PUBLIC HEALTH RELEVANCE: This proposal describes studies to elucidate the molecular mechanisms whereby the sensor transducer protein, BlaR1, facilitates antibiotic resistance in methicillin-resistant Staphylococcus aureus (MRSA), currently a global clinical scourge. The results will advance our abilities to cope with acceleration of multi-drug resistance in MRSA and other bacterial pathogens.

描述（由申请人提供）：耐甲氧西林金黄色葡萄球菌（MRSA）是一种多药耐药细菌病原体，已成为全球临床威胁，占医院缴纳的感染的近三分之一。 MRSA威胁尤其是阴险的，因为它对2-腹膜抗生素的耐药性（例如青霉素，卡贝苯甲，头孢菌素）仍然是诊所中使用最广泛的抗诊所。因此，阐明MRSA耐药性的分子基础是制定其遏制策略的重要性。 MRSA的主要抗药性机制是否会产生水解？-lactamam抗生素。该产生是由跨膜传感器/传感器蛋白Blar1触发的。暴露于？-lactam抗生素触发了blar1的信号转导，该抗生素会导致？ - 乳酰胺酶产生。最近的CD和IR研究表明，信号转导需要在Blar1的细胞外传感器结构域中的构象转变（Blars）在其结合？-lactam抗生素的结合后。在原子水平上定义这些构象转变已成为阐明信号转导机制的努力的关键目标。尽管有高分辨率蛋白质结构的可用性，但驱动Blar1信号转导的构象转变的原子水平机制仍然晦涩难懂。我们已经开始研究Blars，我们的初步结果表明，一组新的分子因素对BLARS构象转变很重要：蛋白质构象动力学。具体而言，使用溶液核磁共振（NMR），我们观察到？-lactam结合会导致局部BLARS灵活性的位点特异性变化。因此，为了阐明BLAR1信号转导的关键事件，我们将研究BLARS构象动力学的功能后果。因此，我们提出的三个特定目标将定义蛋白质动力学和构象变化之间的相互作用如何促进BLARS的信号转导。这些目的包括：（i）通过一系列不同的？-lactam底物比较激活时BLARS灵活性的位点特异性变化； （ii）定义Blars之间的相互作用机理，以及源自Blar1受体的跨膜区域的细胞外环2的肽； （iii）比较非 - ??-乳酸乳酰胺抑制剂与blar1 vets？-lactam底物（抗生素），对Blars的动力学和构象的影响。我们的主要实验工具将是多维NMR，它为蛋白质动力学的原子级描述提供了一种具有独特功能的方法，以抗氨基酸性的基础为基础。我们的研究将为MRSA中负责有害的信号转导机制的内部工作提供新的启示。公共卫生相关性：该提案描述了阐明分子机制的研究，通过这些研究，传感器传感器蛋白BLAR1促进了甲氧西林抗药性金黄色葡萄球菌（MRSA）的抗生素耐药性，目前是一项全球临床冲突。结果将提高我们应对MRSA和其他细菌病原体中多药耐药性加速的能力。