NAMS: Native ambient mass spectrometry

Lead Research Organisation: University of Birmingham
Department Name: Sch of Biosciences

Abstract

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.

Planned Impact

Who will benefit from the research?

The beneficiaries of this research will be analytical instrument manufacturers, the pharmaceutical industry, the NHS and its patients, and the UK's National Measurement Institute (the National Physical Laboratory, NPL).

How will they benefit from this research?

The proposed research has the potential to contribute significantly to the nation's health and wealth:

Analytical instrumentation manufacturers:
The work will enable the development of native ambient mass spectrometry, a set of tools which can be employed in a broad range of new applications. According to InnovateUK1, Measurement Science and Technology plays a crucial role in supporting UK industry and research with an estimated £7 billion turnover in the UK and > 200,000 scientists employed. There are >1700 accredited laboratories and 11,000 sites with analytical laboratories in the UK.

Project partners Waters Corp., Thermo Fisher and Advion Inc. are global companies, each with a strong base in the UK. Owlstone Ltd. is a small medium enterprise, established in 2004. The proposed work will lead to increased sales thus fostering global economic performance and the economic competitiveness of the UK.

National Measurement Institute:
The research will facilitate the UK's national measurement institute (project partners NPL) in achieving its goal of providing 'world-leading measurement solutions which are critical to commercial research and developments and supporting business across the UK and the globe'.

Pharmaceutical industry:
The pharmaceutical industry is underpinned by analytical science. According to the 2016 report on 'Strength & Opportunity in the UK Life Sciences', commissioned by the Office for Life Sciences, the Life Sciences industry employs >200, 000 people in nearly 6000 companies, with an annual turnover of £61 bn. The biopharmaceutical industry employs >100,000 people in 2000 companies generating an annual turnover of £40 bn. The research willovide new analytical tools for determining the effects of drugs on tissue biochemistry, drug-target engagement and efficacy. Project partners include AstraZeneca, a global company with a UK base.

NHS and its patients:
Incidence of Alzheimer's disease is rapidly increasing alongside our ageing population. Approximately 5% of the European population have Alzheimer's2. Worldwide, over 47 million people have Alzheimer's. The cost to the UK of dementia is expected to more than double from £26bn to £55bn by 20403. The tools developed will be applied to mapping and prediction of disease progression through in situ analysis of protein misfolding and aggregation. The research has the potential to improve speed and accuracy of diagnosis by providing a route to understanding the biological changes underpinning Alzheimer's disease.

Incidence of non-alcoholic fatty liver disease (NAFLD) is dramatically increasing alongside increasing obesity rates. Up to 40% of the general population are estimated to have NAFLD 4 with over 60% of adults in England being overweight or obese5. The cost to the NHS is £4.2 bn and that is predicted to double by 20506. NAFLD covers a range of disease states, the most severe of which is non-alcoholic steatohepatitis (NASH). NASH is potentially fatal, leading to cirrhosis, liver failure and cancer. The only treatment option is liver transplantation. There is currently no strategy for predicting disease progress. The proposed research will provide insight into disease progression, thereby enabling improved, and earlier, diagnosis.

1. connect.innovateuk.org/web/3346502
2. Niu, H., Neurologica, 2016. DOI: 10.1016/j.nrl.2016.02.016.
3. www.dementiastatistics.org.
4. LaBrecque, D., World Gastroenterology Organisation Global Guidelines: Nonalcoholic Fatty Liver Disease and Nonalcoholic Steatohepatitis. 2012.
5. digital.nhs.uk/catalogue/PUB19295.
6. www.britishlivertrust.org.uk/about-us/media-centre/facts-about-liver-disease/

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