A microscale thermophoresis instrument for characterizing molecular interactions

近年来，微尺度热泳术（MST）已成为一种强大的技术，用于表征各种分子（如蛋白质、DNA和脂质）之间的相互作用。MST依赖于热泳现象，即分子在温度梯度下的运动。与其他生物物理技术相比，MST有几个优点：1)它非常敏感，需要很少的样品；ii)分析可在复杂缓冲液或样品中进行，如血清或细胞裂解液；iii)数据收集简单，快速，在一次性毛细管中进行，消除污染并允许有效的用户培训。这项资助将为7名nserc资助的共同申请人和他们研究小组中的约43名高素质人员提供微型热泳仪（MST）。共同申请者将研究广泛的生物技术和生物医学问题，并将能够完成特定的目标，如1)表征疏水蛋白与蛋白质和生物材料的相互作用，2)确定黑素细胞发育的调节机制，3)表征分子拥挤对强制外消旋酶抑制剂的影响，4)研究KapB参与细胞质加工体的调节，5)了解FAST蛋白如何与细胞囊泡相互作用以诱导膜融合；6)开发苦味受体抑制剂；7)研究修饰的p53蛋白与细胞结合伙伴的相互作用；8)研究小分子如何影响代谢途径。

Recently, microscale thermophoresis (MST) has emerged as a powerful technique that is used to characterize interactions between a variety of molecules such as proteins, DNA and lipids. MST relies on the phenomenon of thermophoresis, which is the movement of molecules when placed in a temperature gradient. There are several advantages that MST has over other biophysical techniques: i) It is extremely sensitive and requires very little sample; ii) Analysis can take place in complex buffers or samples such blood serum or cell lysate; iii) Data collection is simple, fast and takes place in disposable capillaries, eliminating contamination and allowing for efficient user training. This grant will provide ready access of a microscale thermophoresis (MST) instrument to the 7 NSERC-funded co-applicants and the ~43 highly qualified personnel in their research groups. The co-applicants study a wide range of biotechnical and biomedical problems and will be able to complete specific goals such as 1) characterize hydrophobin interactions with proteins and biomaterials, 2) determine the regulatory mechanisms of melanocyte development, 3) characterize the effect of molecular crowding on mandalate racemease inhibitors 4) investigate the involvement of KapB in the regulation of cytoplasmic processing bodies, 5) gain insight to how FAST proteins interact with cellular vesicles to induce membrane fusion, 6) develop inhibitors to bitter taste receptors, 7) study the interactions of modified p53 proteins with cellular binding partners, and 8) investigate how small molecules can influence metabolic pathways. No similar equipment is available to the applicants, making it essential to acquire a MST instrument to support the applicants’ research programs. The instrument will be located in the Dalhousie University Proteomics Core Facility, allowing for easy access of the instrument to other researchers in the Maritime region. A full time staff member will be responsible for the instrument and providing training to HQP interested in using it. This will allow trainees to learn to conduct MST experiments and interpret results, which will be a valuable skillset in their future careers.