Integrated liquid chromatography-multiangle light scattering system to measure absolute masses and stoichiometries of biological assemblies in solution

集成液相色谱-多角度光散射系统，用于测量溶液中生物组件的绝对质量和化学计量

• 批准号: RTI-2019-00728
• 负责人: Melacini, Giuseppe
• 金额: $ 10.92万
• 依托单位: McMaster University
• 依托单位国家: 加拿大
• 项目类别: Research Tools and Instruments
• 财政年份: 2018
• 资助国家: 加拿大
• 起止时间: 2018-01-01 至 2019-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=659504
• 关键词: Integrated; liquid; chromatography; multiangle; light

This application requests a state-of-the-art size exclusion chromatography system integrated with multi-angle light scattering (SEC-MALS). SEC-MALS is capable of measuring absolute aggregate molecular weights (MWs) and radii of gyration (Rg) under physiological native solution conditions with unparalleled accuracy, precision and throughput not attainable by conventional analytical SEC. Reliable MW and Rg values are needed to determine the oligomerization state and binding stoichiometry of complexes formed by a wide range of proteins and polymers at the center of a multitude of research programs at McMaster. These include not only globular proteins, but also membrane proteins, intrinsically disordered proteins, non-fouling polymers as well as amyloidogenic polypeptides, which are notoriously refractory to other methods for measuring MW and Rg. The requested instrument, a Wyatt MALS-RI with GE Healthcare SEC, is one of the most versatile and reliable SEC-MALS configurations for academic work currently on the market and will have a transformative impact on a collaborative community of > 8 different NSERC-funded research groups spread across three different faculties at McMaster university, i.e. Faculty of Science, Faculty of Health Sciences and Faculty of Engineering. Currently, there is no SEC-MALS at McMaster. Given the outstanding performance in terms of accuracy, sensitivity and throughput, SEC-MALS is now the gold standard requested by the best international journals for the accurate determination of oligomeric states and macromolecular complex stoichiometries. Without SEC-MALS it is becoming nearly impossible to publish high-impact papers. Furthermore, SEC-MALS has been adopted as the industry standard for the characterization of higher-order structures formed by therapeutic biologicals and polymers. Hence, SEC-MALS will dramatically enhance our ability to train the next generation of HQP in preparation for jobs not only in the academia, but also in the pharma and biotech-sectors. The absence of SEC-MALS at McMaster is severely limiting dozens of researchers and is jeopardizing their ability to maintain internationally competitive research programs. If funded, the requested instrument will be the first SEC-MALS in the whole McMaster campus and will be installed in a newly-renovated centralized, shared and widely accessible core facility. We anticipate an extremely high demand by all groups working in the structural biology, enzymology and bioengineering fields. For virtually all projects in these fields, it is imperative to reliably establish the stoichiometry of homo- and hetero-oligomers before further progress can be achieved and currently SEC-MALS is the prime technique for this purpose. The impact of the research enabled by the requested SEC-MALS ranges from the understanding of the mechanisms of allosteric regulation to the design of new biomaterials. Given the absence of SEC-MALS at McMaster, SEC-MALS is a clear and urgent need. **

本申请要求一种集成多角度光散射（SEC-MALS）的最先进的尺寸排阻色谱系统。SEC-MALS能够在生理天然溶液条件下测量绝对聚集体分子量（MW）和回转半径（Rg），具有传统分析SEC无法达到的无与伦比的准确度、精密度和通量。可靠的MW和Rg值需要确定由各种蛋白质和聚合物形成的复合物的低聚状态和结合化学计量，这些蛋白质和聚合物是McMaster众多研究项目的中心。这些不仅包括球状蛋白，而且还包括膜蛋白、固有无序蛋白、无污染聚合物以及淀粉样蛋白生成多肽，众所周知，这些多肽对于用于测量MW和Rg的其他方法是难处理的。所要求的仪器是Wyatt MALS-RI和GE Healthcare SEC，是目前市场上学术工作中最通用和最可靠的SEC-MALS配置之一，将对麦克马斯特大学三个不同学院（即理学院，健康科学学院和工程学院）的超过8个不同NSERC资助的研究小组的协作社区产生变革性影响。 目前，麦克马斯特没有SEC MALS。 由于在准确性、灵敏度和通量方面的出色表现，SEC-MALS现在是最好的国际期刊所要求的准确测定低聚物状态和大分子复合物化学计量的黄金标准。没有SEC-MALS，发表高影响力的论文几乎是不可能的。此外，SEC-MALS已被用作表征治疗性生物制剂和聚合物形成的高阶结构的行业标准。因此，SEC-MALS将大大提高我们培养下一代HQP的能力，不仅为学术界的工作做准备，而且还为制药和生物技术部门做准备。麦克马斯特大学没有SEC-MALS，严重限制了数十名研究人员，并危及他们维持国际竞争力研究项目的能力。 如果获得资助，所要求的仪器将是整个麦克马斯特校园的第一台SEC-MALS，并将安装在一个新装修的集中式，共享和广泛访问的核心设施中。我们预计在结构生物学，酶学和生物工程领域工作的所有团体的需求非常高。 对于这些领域的几乎所有项目，在取得进一步进展之前，必须可靠地建立同源和异源低聚物的化学计量，目前SEC-MALS是用于此目的的主要技术。SEC-MALS所要求的研究的影响范围从对变构调节机制的理解到新生物材料的设计。鉴于麦克马斯特没有SEC-MALS，SEC-MALS是一个明确和迫切的需要。 **