Bacterial back talk - unfolding and molecular interaction studies in Staphylococcal proteins and engineering structure-informed antibiotics

细菌顶嘴——葡萄球菌蛋白和工程结构知情抗生素的展开和分子相互作用研究

• 批准号: 2182232
• 金额: --
• 依托单位: University of Nottingham
• 依托单位国家: 英国
• 项目类别: Studentship
• 财政年份: 2018
• 资助国家: 英国
• 起止时间: 2018 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=studentship-2182232
• 关键词: Bacterial; back; talk; unfolding; molecular

Staphylococci are extremely common human commensals, which have been implicated in systemic infections in hospitals, biofilm formation in surgical devices and implants, industrial fouling and food spoilage. The accessory gene regulation (Agr) in staphylococci controls important bacterial functions, such as virulence and adhesion to surfaces (biofilm formation). AgrB is a membrane endopeptidase responsible for pre-processing of the signalling molecule, AIP, which mediates bacterial communications and quorum sensing. The AIP response cascade is a two component system including a membrane receptor AgrC, which phosphorylates the nucleotide-binding partner AgrA. Pharmacological regulation of the Agr protein is an attractive target for bacterial control, attenuating virulence and biofilm inhibition and a new opportunity for antimicrobial design.We have developed a new molecular recognition assay, which reports on substrate or ligand binding through increased protein stability to degradation. Thermal unfolding of proteins is a gradual process, in which thermal fluctuations increase in frequency and magnitude with temperature and can be monitored by circular dichroism, small angle neutron or X-ray scattering. NMR spectroscopy and high performance computing complement the studies with detailed atomic level structural information on the interaction between the Agr proteins and their substrates and regulatory ligands. In this project we will develop a large scale screen for protein/ligand interactions based on the thermal denaturation assay and will validate it against existing libraries of Agr modulators. Using recombinant protein from current research project on Agr, we will:a) use a combination of scattering techniques and CD to monitor changes in protein structure and overall conformation and will develop the tool with reference to critical behavior;b) develop a sample environment at partner institution ISIS for the large scale screen in conjunction with beamlines SANS2D and ZOOM; develop analytical framework for rapid data interpretation;c) characterize ligand/receptor interactions using solid state NMR, molecular docking and simulations.The programme relies on advanced biophysical, structural and computational tools for molecular analysis, which are combined in complementary, yet partially overlapping groups to ensure successful completion of the work.

葡萄球菌是极为常见的人类份量，与医院的全身感染，外科手术装置和植入物，工业结垢和食物变质有关。葡萄球菌中的辅助基因调节（AGR）控制着重要的细菌功能，例如毒力和对表面的粘附（生物膜形成）。 AGRB是一种负责预处理信号分子AIP的膜内肽酶，它介导细菌通信和法规传感。 AIP响应级联反应是一个两个组件系统，其中包括膜受体AGRC，它磷酸化核苷酸结合伴侣Agra。 AGR蛋白的药理调节是细菌控制，衰减毒力和生物膜抑制的有吸引力的靶标，以及用于抗菌设计的新机会。我们开发了一种新的分子识别测定法，该测定法报告了底物或配体通过增加蛋白质稳定性降解的底物或配体结合。蛋白质的热展开是一个逐渐的过程，其中热波动随温度而增加的频率和幅度增加，可以通过圆形二色性，小角度中子或X射线散射来监测。 NMR光谱和高性能计算补充了有关AGR蛋白及其底物和调节配体之间相互作用的详细原子水平结构信息。在这个项目中，我们将根据热变性测定法开发一个大规模筛选，以针对蛋白质/配体相互作用进行大规模筛选，并将针对AGR调节剂的现有库进行验证。我们将使用当前有关AGR研究项目的重组蛋白，我们将：a）使用散射技术和CD的组合来监测蛋白质结构的变化和整体构象的变化，并会参考关键行为来开发该工具； b）在与Beamlines SANS2D和Zoom结合的大型筛选中，在伙伴ISIS开发了ISIS的样本ISIS；开发用于快速数据解释的分析框架； c）使用固态NMR，分子对接和模拟表征配体/受体相互作用。该程序依赖于先进的生物物理，结构和计算工具用于分子分析，这些工具与互补但部分重叠的组结合在一起，以确保完成工作的完成。