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用于确定生物分子相互作用和描述溶液中蛋白质稳定性的尖端生物物理工具

Cutting edge biophysical tools to determine biomolecular interactions and characterize protein stability in solution
课程网址: http://videolectures.net/biophysics2018_nowak_biophysical_tools/  
主讲教师: Jakub Nowak
开课单位: 纳米热处理技术有限公司
开课时间: 2018-07-09
课程语种: 英语
中文简介:
介绍蛋白质-蛋白质、蛋白质核酸或蛋白质小分子等生物分子相互作用的微尺度热泳分析,不仅有助于发展治疗或诊断技术,而且还提供了对细胞过程的重要见解。在这里,我们提出了微尺度热泳(MST)的研究亲和力的生物分子相互作用。MST是一种生物物理技术,通过检测红外激光引起的温度变化引起的荧光信号变化来测量两个分子之间相互作用的强度。荧光信号的变化范围与配体与荧光靶的结合有关。纳米DSF中蛋白质中色氨酸的荧光强烈依赖于其周围环境。通过跟踪荧光、化学和热稳定性的变化,可以以真正无标签的方式进行评估。NanoTemper的双UV技术允许快速蛋白质展开分析,提供无与伦比的扫描速度和数据点密度。这就产生了超高分辨率的蛋白质熔融图谱,允许检测甚至微小的去折叠信号。此外,由于不需要二级报告荧光团,因此可以独立于缓冲液成分分析蛋白质溶液。此外,可以并行记录蛋白质聚集的信息,从而深入了解样品的胶体稳定性。本演示将涵盖该技术的生物物理概念,展示MST和nanoDSF技术平台的优势,随后将介绍各种实验系统的具体应用示例。
课程简介: Introduction to Micro Scale Thermophoresis Analysis of bio-molecular interactions, such as protein-protein, protein-nucleic acid or protein-small molecule, not only helps to develop therapeutics or diagnostics techniques, but it also provides important insights into cellular processes. Here we present MicroScale Thermophoresis (MST) for the investigation of affinities of biomolecular interactions. MST is a biophysical technique that measures the strength of the interaction between two molecules by detecting variations in fluorescence signal as a result of an IR-laser induced temperature change. The range of the variation in the fluorescence signal correlates with the binding of a ligand to the fluorescent target Introduction to nanoDSF The fluorescence of tryptophans in a protein is strongly dependent on its close surroundings. By following changes in fluorescence, chemical and thermal stability can be assessed in a truly label-free fashion. The dual-UV technology by NanoTemper allows for rapid protein unfolding analysis, providing an unmatched scanning speed and data point density. This yields ultra-high resolution melting profiles of proteins which allow for detection of even minute unfolding signals. Furthermore, since no secondary reporter fluorophores are required, protein solutions can be analyzed independent of buffer compositions. In addition, information on protein aggregation can be recorded in parallel, providing insight into colloidal stability of the sample. The presentation will cover biophysical concepts of the techniques showing benefits of MST and nanoDSF technologies platforms, and will be followed by specific examples of applications towards various experimental systems.
关 键 词: 生物分子; 生物物理技术; 微尺度热泳分析
课程来源: 视频讲座网
数据采集: 2021-11-12:zkj
最后编审: 2021-11-12:zkj
阅读次数: 75