DETECTION OF ACH-MEDIATED CONFORMATIONAL CHANGES OF ACHBP

ACH介导的ACHBP构象变化的检测

• 批准号: 7598788
• 负责人: Steven M Sine
• 金额: $ 0.2万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-03-01 至 2008-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7598788
• 关键词: Agonist; Binding Sites; Chemicals; Cholinergic Receptors; Computer Retrieval of Information on Scientific Projects Database; Cysteine; Detection; Family; Funding; Gated Ion Channel; Grant; Institution; Label; Ligand Binding Domain; Ligands; Mediating; Molecular Conformation; Mutate; Peripheral; Proteins; Research; Research Personnel; Resources; Serine; Source; United States National Institutes of Health; member; molecular dynamics; mutant

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Acetylcholine receptor (AChR) is a member of ligand-gated ion channel family. Small agonists like ACh trigger the channel opening by changing its conformation from the closed state to the open state. Here we want to study ACh-mediated conformational changes to AChBP, a soluble protein that is homologous to the ligand binding domain of AChR. Our molecular dynamics simulation (JBC in press) indicates ACh triggers the collapse of a peripheral loop region called C-loop on the binding site. At the tip of the C-loop, two vicinal cysteines have dramatic conformational changes. We isotopically-labeled only cysteine residues (U-13C3, 15N) in the protein and plan to see change of chemical shift in NMR spectra following addition of ACh. Since there are other cysteines in the protein, the spectra of a mutant protein with the vicinal cyteines mutated to Serine will be compared.

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 乙酰胆碱受体(AChR)是配体门控离子通道家族的一员。像ACh这样的小激动剂通过将其构象从关闭状态改变为开放状态来触发通道开放。在这里，我们想要研究ACh介导的AChBP的构象变化，AChBP是一种与AChR的配体结合域同源的可溶性蛋白质。我们的分子动力学模拟(JBC在出版)表明，ACh触发了结合位点上名为C-loop的外围环区的崩溃。在C环的末端，两个邻近的半胱氨酸发生了显著的构象变化。我们只对蛋白质中的半胱氨酸残基(U-13C3，15N)进行了同位素标记，并计划在添加ACh后看到核磁共振谱中化学位移的变化。由于蛋白质中还有其他半胱氨酸，因此我们将比较突变蛋白的光谱，其中邻近的半胱氨酸突变为丝氨酸。