Membrane Protein Stability

膜蛋白稳定性

• 批准号: 7103700
• 负责人: JAMES U BOWIE
• 金额: $ 27.26万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-08-01 至 2009-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7103700
• 关键词: Escherichia coli; Halobacteriaceae; X ray crystallography; bacterial genetics; bacteriorhodopsins; calorimetry; chemical kinetics; chemical stability; crystallization; enzyme induction /repression; gene mutation; hydrogen bond; mass spectrometry; membrane proteins; model design /development; permease; phosphotransferases; physical model; point mutation; protein folding; protein structure function; racemization; solubility; structural biology; thermodynamics

DESCRIPTION (provided by applicant): Knowledge of energetic contributions to protein folding is essential for many of the central quests of structural biology, including protein design and modeling, as well as an understanding of protein function and dysfunction in disease. Consequently, an enormous effort has been made to work out the forces that stabilize protein structure. In this effort, membrane proteins have been largely neglected, in spite of the fact that they play critical biological roles and are the major targets of drugs. Membrane protein folding has been avoided because of the enormous complexity of the membrane environment and the technical challenges faced when studying this problem. In the prior grant period, we have overcome many of these challenges and are finally well positioned to dissect the energetics of folding in a large membrane protein. To my knowledge, there is no other group that combines both thermodynamic stability measurements and detailed structural analysis in a similar way. The Specific Aims are: I.To probe packing forces, we will introduce bumps and create holes at various points in the bacteriorhodopsin (bR) structure and examine the consequences for bR structure and stability. II. To probe hydrogen bond strength, we will specifically delete inter-helical hydrogen bonds in bR and examine the consequences for bR structure and stability. III. To probe the contribution of interhelical loops, we will examine the consequences of side chain deletions and loop insertions on bR structure and stability. IV. Develop hydrogen exchange as a means to probe the unfolded state of bR.

描述（由申请人提供）：了解蛋白质折叠的能量贡献对于结构生物学的许多核心任务至关重要，包括蛋白质设计和建模，以及对疾病中蛋白质功能和功能障碍的理解。因此，人们付出了巨大的努力来找出稳定蛋白质结构的力。在这一努力中，膜蛋白在很大程度上被忽视，尽管它们发挥着关键的生物学作用并且是药物的主要靶标。由于膜环境的巨大复杂性以及研究该问题时面临的技术挑战，膜蛋白折叠已被避免。在之前的资助期间，我们已经克服了许多这些挑战，并最终有能力剖析大膜蛋白折叠的能量学。据我所知，没有其他小组以类似的方式将热力学稳定性测量和详细的结构分析结合起来。 具体目标是： I. 为了探测堆积力，我们将在细菌视紫红质 (bR) 结构的各个点引入凸块并创建孔，并检查对 bR 结构和稳定性的影响。 二.为了探测氢键强度，我们将专门删除 bR 中的螺旋间氢键，并检查对 bR 结构和稳定性的影响。 三．为了探讨螺旋间环的贡献，我们将检查侧链删除和环插入对 bR 结构和稳定性的影响。 四．开发氢交换作为探测 bR 未折叠状态的一种手段。