NAMS: Native ambient mass spectrometry

这是该奖学金的延伸：“NISA：生物分子原位分析的新方法”（EP/L023490/1）。原始研究的目的是开发原位生物分子分析的新方法，即，直接从自然（或实际）环境中分析生物分子。主要的重点是蛋白质的原位分析。蛋白质是细胞的工作马，执行生命所需的所有功能。它们还被用作治疗剂和消费品。为了深入了解蛋白质在生命过程中的作用，有必要在分子水平上分析蛋白质。质谱法根据其质荷比表征电离分子，非常适合这一挑战，提供高灵敏度，广泛的特异性（所有分子都有质量）和化学结构解析能力。最初的奖学金中的大部分研究集中在开发用于原位分析完整但未折叠的蛋白质的质谱工具。通过质谱与气相分离技术的联用和质谱仪器的改进，灵敏度已取得重大进展。这些工具能够识别未知蛋白质，识别和定位蛋白质修饰或突变的位点，以及底物内蛋白质的空间分布（质谱成像）。然而，这些工具并不能提供蛋白质整体三维结构的信息。蛋白质的三维结构决定了它们的功能。因此，蛋白质结构的知识对于破译蛋白质在健康和疾病中的作用至关重要。为了充分询问蛋白质的结构，功能和环境之间的关系，有必要开发工具，结合天然质谱，其中蛋白质保持在其折叠形式和它们的分子间和分子内的非共价相互作用的维持。为了满足这一需求，作为原始奖学金的一部分进行的初步研究集中在开发天然环境质谱法，其中折叠蛋白质，蛋白质复合物和蛋白质组装体直接从其生理环境中取样。到目前为止，我们在这一领域的研究集中在一个单一的采样技术，即，液体萃取表面分析;然而，有许多环境采样方法可能被证明是合适的，每种方法在灵敏度、速度和空间分辨率方面提供不同的规格。奖学金延期的目的是建立自然环境质谱法作为一门广泛的学科折叠蛋白质及其复合物的原位分析。目标是开发一套工具，能够提供有关蛋白质在健康和疾病中的功能的信息。原生环境质谱法的每一种潜在应用都将带来其独特的挑战。例如，空间分辨率，即，组织中蛋白质分布的复杂映射可能是至关重要的要求。或者，高通量（分析速度）可能是成功的关键，或者可能是技术的灵敏度至关重要。通过扩大原生环境质谱的范围，以涵盖全方位的采样技术，我们将能够解决这些挑战中的每一个。此外，一系列的离子迁移谱技术，使蛋白质结构的测量以及提高灵敏度，将与本地环境质谱允许三维蛋白质结构的空间分布集成。这项研究的影响将通过应用于阿尔茨海默病（一种与蛋白质错误折叠和聚集有关的疾病）和非酒精性脂肪肝（一种与蛋白质和脂质之间异常结合有关的疾病）来证明。

This is an extension of the Fellowship: 'NISA: Novel approaches for in situ analysis of biomolecules' (EP/L023490/1).The aim of the original research was to develop novel approaches for in situ biomolecular analysis, i.e., the analysis of biomolecules directly from their natural (or actual) environment. The principal focus has been on the in situ analysis of proteins. Proteins are the work-horses of the cell and perform all the functions required for life. They also find uses as therapeutics and in consumer products. To gain insight into the roles of proteins in life processes, it is necessary to analyse proteins at a molecular level. Mass spectrometry, in which ionised molecules are characterised according to their mass-to-charge, is ideally suited to this challenge, offering high sensitivity, broad specificity (all molecules have a mass), and the capability for chemical structure elucidation. The majority of research within the original fellowship has concentrated on development of mass spectrometry tools for in situ analysis of INTACT, but UNFOLDED, proteins. Significant advances in sensitivity have been achieved through hyphenation of mass spectrometry with gas-phase separation techniques and modifications to the mass spectrometry instrumentation. These tools enable identification of unknown proteins, identification and localisation of sites of protein modification or mutation, and spatial profiling (mass spectrometry imaging) of proteins within the substrate. Those tools do not, however, provide information on the overall 3-D structure of proteins. It is the 3-D structure of proteins that dictate their function. Knowledge of protein structure is therefore vital in deciphering the roles of protein in health and disease. In order to fully interrogate the relationship between protein structure, function and environment, it is necessary to develop tools incorporating native mass spectrometry in which proteins remain in their FOLDED form and their inter- and intra-molecular noncovalent interactions are maintained. To address that need, preliminary research undertaken as part of the original fellowship has focused on developing methods for NATIVE AMBIENT MASS SPECTROMETRY in which folded proteins, protein complexes and protein assemblies are sampled directly from their physiological environment. To date, our research in this area has focused on a single sampling technique, i.e., liquid extraction surface analysis; however, there are many ambient sampling approaches which may prove suitable, each offering different specifications in terms of sensitivity, speed, and spatial resolution.The aim of the fellowship extension is to establish NATIVE AMBIENT MASS SPECTROMETRY as a broad discipline for the in situ analysis of folded proteins and their complexes. The goal is to develop a suite of tools which will be capable of providing information on protein function in health and disease. Each potential application of native ambient mass spectrometry will come with its own unique challenges. For example, spatial resolution i.e., intricate mapping of the protein distribution in the tissue, may be the crucial requirement. Alternatively, high throughput (speed of analysis) may be the key to success, or it may be that the sensitivity of the technique that is vital. By widening the scope of native ambient mass spectrometry to encompass a full range of sampling techniques, we will enable each of these challenges to be addressed. Moreover, a range of ion mobility spectrometry techniques, which enable measurement of protein structure as well as improving sensitivity, will be integrated with native ambient mass spectrometry allowing spatial profiling of 3D protein structure. The impact of the research will be demonstrated by application to Alzheimer's disease, a disease associated with protein misfolding and aggregation, and non-alcoholic fatty liver disease, a disease associated with unusual binding between proteins and lipids.