CHARACTERIZATION OF NOVEL LIPID CUBIC PHASE MATRICES FOR MEMBRANE PROTEIN CRYSTA

膜蛋白晶体新型脂质立方相基质的表征

• 批准号: 8169941
• 负责人: HIROTSUGU TSURUTA
• 金额: $ 0.24万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-05-01 至 2011-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8169941
• 关键词: Architecture; Behavior; Computer Retrieval of Information on Scientific Projects Database; Crystallization; Detergents; Drops; Film; Funding; G-Protein-Coupled Receptors; Grant; Human; Institution; Joints; Lipids; Measurement; Membrane; Membrane Proteins; Phase; Property; Proteins; Research; Research Personnel; Resolution; Resources; Sampling; Solvents; Source; Techniques; Technology; Temperature; United States National Institutes of Health; design; innovation; novel

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. High-resolution structural studies of membrane proteins are limited by the availability of crystals diffracting to high resolution. Crystallization in lipidic cubic phase (LCP) matrices (or in meso) has proven to yield high quality crystals of challenging membrane proteins, such as human G protein-coupled receptors. Broader applications of in meso techniques require identification of new lipids with specific phase properties capable of stabilizing proteins with a wide range of sizes and architectures. The Joint Center for Innovative Membrane Protein Technologies at TSRI designs and synthesizes such lipids. The phase and structural behavior of novel lipid matrices should be thoroughly characterized prior to being used in specific applications. JCIMPT has developed the LCP sandwich plates in which small drops of lipid-solvent-protein mixture are held between two x-ray transparent synthetic films with a thin spacer to form 8x12 grids of samples similar to the standard 96-well microplate format for membrane crystallization. The BTP staff has built a temperature controlled holder for the LCP sandwich plate and conducted first trial measurements on BL4-2. One of the two synthetic films was found superior to the other for x-ray scattering studies, providing a low-level of reasonably flat scattering background. We believe our approach will allow us to study effects of detergents, additive lipids, proteins as well as great variety of precipitants on the lipidic matrices in a high-throughput mode under the conditions mimicking those encountered during membrane protein crystallization trials.

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 膜蛋白的高分辨率结构研究受到衍射到高分辨率的晶体的可用性的限制。在介晶立方相（LCP）基质（或内消旋）中结晶已被证明产生具有挑战性的膜蛋白（例如人G蛋白偶联受体）的高质量晶体。更广泛的应用，在介观技术需要识别新的脂质与特定的相特性，能够稳定蛋白质的范围广泛的大小和结构。TSRI的创新膜蛋白技术联合中心设计和合成这种脂质。新型脂质基质的相和结构行为在用于特定应用之前应彻底表征。JCIMPT开发了LCP夹心板，其中小滴的脂质-溶剂-蛋白质混合物被保持在两个X射线透明的合成膜之间，具有薄的间隔物，以形成类似于用于膜结晶的标准96孔微孔板格式的8x 12样品网格。 BTP工作人员为LCP夹层板构建了一个温控保持器，并对BL 4 -2进行了首次试验测量。发现两种合成膜中的一种在X射线散射研究中比另一种优越上级，提供了低水平的合理平坦的散射背景。我们相信，我们的方法将使我们能够研究洗涤剂，添加剂脂质，蛋白质以及各种各样的沉淀剂在高通量模式下，在模拟膜蛋白质结晶试验过程中遇到的条件下，对蛋白质基质的影响。