Applications of NMR Spectroscopy to Study Structure, Dynamics and Small Molecule Interactions Related to Protein Folding and Misfolding

核磁共振波谱是研究自然环境中生物大分子在溶液中的结构、动力学和相互作用的有力工具，包括细胞环境中的蛋白质，以及包括生物膜、生物材料和膜蛋白质在内的固态生物大分子。这种方法使研究人员能够获得这些组分的详细原子分辨率图像，并研究这些生物分子引起的相互作用的强度和频率，给出感兴趣的系统的全面图景。溶液生物分子核磁共振的一个独特特征，特别是最近发展起来的TXO低温探针，是能够表征缺乏明显三维结构的蛋白质的残余结构和瞬时相互作用，这些蛋白质被称为内在无序。真核基因组中约30%的蛋白质以内源性紊乱为主要特征，这些蛋白质在许多细胞过程以及包括癌症、神经退行性疾病、扩张性心肌病在内的人类异常疾病中起着中心作用。生物分子核磁共振的另一个关键研究领域是固态；在固态中，样品由于自组装或与生物膜或蛋白质网络的相互作用而具有更严格的构象。生物分子单核磁共振一直在推动蛋白质淀粉样纤维和膜蛋白的表征，是一种非常精确的方法来探测生物分子组件的结构特性，即生物材料、大的蛋白质复合体和水凝胶。化学系的核磁共振设施一直在为剑桥地区的一大批分子科学家提供服务，包括化学系、错误折叠疾病中心(CMD)和一些公司和其他部门。申请者的研究实验室最近的研究进展使蛋白质聚集和错误折叠疾病领域有了重大发现，为此，CMD获得了资金，以促进寻找治疗包括阿尔茨海默氏症和帕金森氏症在内的神经退行性疾病的过渡研究。CMD以及访问我们的核磁共振设施的其他研究实验室的活动将极大地受益于高通量筛选与这些疾病有关的前体蛋白的小分子。尽管我们的旗舰700 MHz仪器配备了低温探头TXO和样品交换器，但目前这种方法无法有效地应用，我们需要更现代化的光谱仪控制台来提高灵敏度和应用快速数据采集方法。更换控制台是我们更新700 MHz光谱仪的更大计划的一部分，包括对磁铁的维护，以确保更具成本效益的仪器管理并避免未来的故障。除了我们在溶液核磁共振方面的专业知识外，我们还在研究与阿尔茨海默氏症和帕金森病相关的聚集物种方面获得了国际上相关的专业知识。在这种情况下，我们希望为我们的核磁共振设施配备生物固体核磁共振能力，用于蛋白质研究，这在剑桥地区令人惊讶地缺乏。该设备还将允许对与阿尔茨海默氏症和帕金森氏症相关的特定蛋白质聚集体进行药物筛选。除了这一应用将对与研究神经退行性疾病中固有的无序蛋白质和蛋白质聚集相关的CMD活动产生巨大好处外，新的核磁共振能力还将服务于剑桥大学更大的科学家社区和经常访问该设施的合作机构，包括工业合作者，促进和影响涵盖有机化学、材料科学和化学生物学各个方面的研究。

NMR spectroscopy is a powerful technique to study the structure, dynamics and interactions of biomacromolecules in their natural environment in solution, including proteins the cellular milieu, and in the solid-state, including biological membranes, biomaterials and membrane-proteins. This methodology allows researchers to gain detailed atomic resolution pictures of these components and to also study the strength and frequency of the interactions incurred by these biomolecules, giving a comprehensive picture of the systems of interest. A unique feature of the solution biomolecular NMR, in particular recently developed TXO cryoprobes, is the ability to characterise residual structure and transient interactions by proteins which lack distinct three dimensional structures and which are referred to as intrinsically disordered. Intrinsic disorder is a primary characteristic of ~30% of the proteins encoded in the eukaryotic genome and these proteins are central to a number of cellular processes as well as aberrant human diseases including cancer, neurodegenerative disorders, dilated cardiomyopathies. Another key research area in biomolecular NMR is within the solid-state; where samples are in more rigid conformations due to self-assembly or interactions with biological membranes or protein networks. Biomolecular ssNMR has been driving the characterisation of protein amyloid fibrils and membrane proteins, and is an extremely accurate method to probe the structural properties of biomolecular assemblies, i.e.biomaterials, large protein complexes and hydrogels. The NMR facility in the Department of Chemistry has been serving a large community of molecular scientists in the Cambridge area, including the Departments of Chemistry, the Centre for Misfolding Disease (CMD) and a number of companies and other Departments. Recent research developments in the research labs of the applicants have enabled significant discoveries in the field of protein aggregation and misfolding diseases, for which the CMD has been funded to boost the transitional research into finding cures for neurodegenerative disorders include Alzheimer's and Parkinson's diseases. The activities of the CMD, as well as of the other research labs accessing our NMR facility, would greatly benefit from the availability of high-throughput screening of small molecules on the precursor proteins linked with these diseases. This methodology cannot, currently, be applied with efficiency, despite our flagship 700 MHz instrument being equipped with a cryoprobe TXO and sample exchanger, and we need a more modern spectrometer console to boost sensitivity and apply fast methods of data acquisitions. The change of console is part of our bigger plan to rejuvenate our 700 MHz spectrometer, including maintenance to the magnet to ensure more cost effective management of the instrument and avoid future breakage.In addition to our expertise in solution NMR, we have gained internationally relevant expertise in studying the aggregated species that are associated with Alzheimer's and Parkinson's disorders. In this context, we would like to equip our NMR facility with bio-solid NMR capability for protein investigations, which is surprisingly missing in the Cambridge area. This equipment will allow also, the drug-screening of selected molecules to specific protein aggregates associated with Alzheimer's and Parkinson's. In addition to the great benefits that this application would have on the CMD activities associated with the study of intrinsically disordered proteins and protein aggregation in neurodegenerative disorders, the new NMR capability will serve the larger community of scientists at the University of Cambridge and the collaborating institutions that frequently access this facility, including industrial collaborators, faciliting and impacting on research encompassing various aspects of organic chemistry,material sciences and chemical biology.