TrHbs: Biophysical Consequences of a Nonpolar Tunnel

TrHbs：非极隧道的生物物理后果

• 批准号: 6917946
• 负责人: JOEL M FRIEDMAN
• 金额: $ 37.58万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-07-15 至 2008-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6917946
• 关键词: Mycobacterium tuberculosis; X ray crystallography; bacterial proteins; binding sites; biophysics; conformation; genetic recombination; heme; hemoprotein; hydrogen bond; interferometry; intermolecular interaction; ligands; molecular polarity; mutant; nuclear magnetic resonance spectroscopy; photolysis; protein structure function; site directed mutagenesis; spectrometry

DESCRIPTION (provided by applicant): Truncated hemoglobins (trHbs) are a recently discovered class of small oxygen-binding protohemeproteins, widely distributed in prokaryotes. Many occur in bacteria pathogenic to man (e.g. Mycobacterium tuberculosis (TB), Mycobacterium avium, Mycobacterium leprae, Cornybacterium diphtheriae, Bordetella pertussis, Legionella pneumophila, Staphylococcus aureus and Bacillus anthracis). This project will focus on trHbN and trHbO, the two trHbs from TB. TrHbN protects aerobic respiration from the inhibitory action of nitric oxide and enhances infectivity. The function oftrHbO is essential for bacterial growth. Since trHbs are not present in man, this makes them potential drug targets. The emphasis of this project is on probing those unusual biophysical properties of these two trHbs that are hypothesized to be the basis for the unusual functionalities that are of potential biomedical significance. Several specific aims are designed to expose the nature and significance of the hydrogen bonding network found in the distal hempocket and of the large apolar tunnel that links the solvent to the ligand binding site at the heme. To achieve in depth insight into how the hydrogen bonding network and the apolar tunnel functioning of these two important proteins, several techniques will be used including photolysis, stopped-flow spectrometry, vibrational spectroscopy (FTIR, UV and visible resonance Raman), structure analysis (X-ray diffraction, NMR), site directed mutagenesis and conformational trapping and kinetics of ligand rebinding. In parallel with this part of the overall project, genetic and molecular biological approaches will be used to correlate the biophysical results with the in vivo functional studies.

描述（由申请人提供）：截短血红蛋白（trHb）是最近发现的一类小的氧结合原血红素蛋白，广泛分布于原核生物中。许多存在于对人类致病的细菌中（例如结核分枝杆菌（TB）、鸟分枝杆菌、麻风分枝杆菌、白喉棒状杆菌、百日咳杆菌、嗜肺军团菌、金黄色葡萄球菌和炭疽杆菌）。该项目将重点关注来自TB的两种trHb--trHbN和trHbO。TrHbN保护有氧呼吸免受一氧化氮的抑制作用，并增强感染性。trHbO的功能是细菌生长所必需的。由于trHbs不存在于人体中，这使它们成为潜在的药物靶点。该项目的重点是探索这两种trHbs的不寻常的生物物理特性，这些特性被假设为具有潜在生物医学意义的不寻常功能的基础。几个具体的目的是暴露的性质和意义的氢键网络中发现的远端hempocket和大的非极性隧道，连接溶剂的配体结合位点的血红素。为了深入了解这两种重要蛋白质的氢键网络和非极性隧道功能，将使用几种技术，包括光解，停流光谱法，振动光谱法（FTIR，UV和可见光共振拉曼），结构分析（X射线衍射，NMR），定点诱变和构象捕获和配体再结合动力学。在整个项目的这一部分的同时，将使用遗传和分子生物学方法将生物物理结果与体内功能研究相关联。