Studies of macromolecular crowding

大分子拥挤的研究

• 批准号: 8349671
• 负责人: Allen P Minton
• 金额: $ 30.77万
• 依托单位: NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8349671
• 关键词: Affinity; Behavior; Binding; Binding Sites; Biological; Cleaved cell; Collaborations; Complex; Crowding; Elements; Enzymes; Fluorescence Resonance Energy Transfer; Goals; Individual; Ligand Binding; Ligands; Liquid substance; Location; Methods; Modeling; Molecular Conformation; Nucleotides; Peptides; Publishing; Relative (related person); Reporting; Site-Directed Mutagenesis; System; Thermodynamics; Time; Work; adenylate kinase; analog; base; fluorophore; macromolecule; theories; trimethyloxamine

In collaboration with the group of E. Haas we have investigated the effect of a small "inert" cosolute, trimethylamine oxide, on the conformational isomerization of an enzyme, adenylate kinase, that is known to undergo significant conformational changes during its catalytic cycle. Binding of substrate nucleotides (AMP, ADP, ATP, or analogs thereof) is accompanied by compaction resulting from the closure of clefts containing ligand binding sites. Conformational changes are being characterized via time-dependent fluorescence resonance energy transfer (FRET) between donor and acceptor fluorophores placed at selected locations along the peptide chain through site-specific mutagenesis. Analysis of the results obtained to date indicate that sufficient concentrations of TMAO stabilize compact or "closed" conformations relative to less compact or "open" conformations, leading to a substantial increase in the affinity of the enzyme for ligands. A report of this work was recently published.

与E.哈斯，我们已经调查了一个小的“惰性”共溶质，氧化三甲胺，对构象异构化的酶，腺苷酸激酶，这是众所周知的，在其催化循环过程中经历显着的构象变化的影响。 底物核苷酸（AMP、ADP、ATP或其类似物）的结合伴随着由含有配体结合位点的裂缝的闭合引起的压实。 构象变化的特点是通过时间依赖性荧光共振能量转移（FRET）之间的供体和受体荧光团放置在选定的位置沿着肽链通过位点特异性诱变。 迄今为止获得的结果的分析表明，足够浓度的TMAO稳定紧凑或“封闭”构象相对于不太紧凑或“开放”构象，导致酶对配体的亲和力的大幅增加。 最近发表了一份关于这项工作的报告。