Analytical Ultracentrifuge

描述(由申请人提供)：托马斯·杰斐逊大学基梅尔癌症中心的关键数量的研究人员有一些悬而未决的项目，这些项目需要一种定量方法来测量生物样本的四级结构及其结合常数。为此，我们建议购买Beckman Optima XL-I分析性超速离心机。该仪器非常适合这些项目，并显著扩展了我们的分子相互作用核心设施的能力。我们将立即利用这一仪器解决的具体领域包括：1)与动力蛋白活性和货物结合调节有关的多聚体动力蛋白和动力蛋白复合体的热力学表征；2)研究与膜融合相关的病毒糖蛋白多肽片段的寡聚状态和流体力学性质；3)tRNA-蛋白质复合体和解码遗传信息的机制；4)鉴定蛋白质相互作用的拮抗剂；5)表征由激活arrestins引起的构象变化；6)表征TCR、pMHC和CD8的外膜结构域的平衡结合常数和化学计量。此外，该仪器将加强另外两个核心设施(X射线和小分子筛查设施)以及研究生教育(结构生物学研究生课程PR613的结构，使学生将教授的每项技术应用于实际样本)。选择的分析超速离心机是Beckman Optima XL-I。该仪器允许使用干涉光学或UV/可见光光学进行检测。干涉模式对折射率很敏感，因此不需要分析物具有天然的色团。因此，这将允许测量具有低分子消光系数的样品、小分子拮抗剂和膜结合蛋白。干涉光学系统还提供了快速数据收集，可用于产生用于沉降速度分析的高质量数据。另一方面，在大范围(190至800 nm)测量吸光度的能力允许同时测量多组分系统中的单个组分。

DESCRIPTION (provided by applicant): A critical number of investigators at the Kimmel Cancer Center of Thomas Jefferson University have a number of pending projects that require a quantitative method for measuring the quaternary structure of biological samples and their association constants. To this end, we propose to acquire the Beckman Optima XL-I analytical ultracentrifuge. This instrument is well suited for these projects and significantly extends the capabilities of our molecular interaction core facility. Specific areas that we will immediately address using this instrumentation are: 1) Thermodynamic characterization of multimeric dynein and dynactin complexes in relation to the regulation of dynein activity and cargo binding; 2) To study the oligomerization state and hydrodynamic properties of peptide segments derived from viral glycoproteins involved in membrane fusion; 3) tRNA-protein complexes and the mechanism of decoding genetic information; 4) Characterization of Protein-Protein Interaction Antagonists; 5) Characterize conformational changes induced by activation of arrestins, a family of proteins that regulate G protein-coupled receptor signaling; 6) Characterization of the equilibrium binding constants and the stochiometry of the ectodomains of the TCR, the pMHC and the CD8. In addition, the instrument will enhance two additional core facilities (X-ray and small molecule screening facilities) as well as graduate education (PR613, a graduate course on structural biology, is structured such that students apply each technique taught to an actual sample). The analytical ultracentrifuge of choice is the Beckman Optima XL-I. This instrument permits detection using either interference optics or UV/visible optics. The interference mode is sensitive to the refractive index, and thus, does not require the analyte to have a natural chromaphor. Thus, this will permit measurement of samples with low molecular extinction coefficients, small molecule antagonist and membrane bound proteins. The interference optics also provides rapid data collection useful for producing high quality data for sedmentation velocity analysis. On the other hand, the ability to measure absorbance over a large range (190 to 800 nM) allows simultaneous measurement of individual components in a multicomponent system.