MUSERMET: MultiUSER equipment for small molecule identification in untargeted METabolomics

Lead Research Organisation: University of Liverpool
Department Name: Institute of Integrative Biology

Abstract

Many of the projects in the newly formed Liverpool Centre for Metabolomics Research involve a requirement for the sophisticated, detailed, and RAPID analysis of biological samples to identify and measure the many hundreds or thousands of small molecules (metabolites) that they contain. This is done by separating them in a flowing stream (liquid chromatography) followed by their estimation on the basis of their masses and the masses of any fragments that may be produced therefrom (as these form a unique 'signature'). Classically, a major bottleneck has been the IDENTIFICATION of the metabolites represented by what may otherwise be highly reproducible times of the appearance of these mass spectral peaks or signatures. An instrument called the Orbitrap ID-X tribrid, together with an integrated suite of software (to which we shall add), offers a real chance of solving this 'spectrum-to-structure' problem. It may be applied to the solution of multiple problems within the BBSRC remit, and we illustrate several.

The aim of this proposal is to secure such an instrument for an extensive User Community in Liverpool and elsewhere. The instrument, while based on established Orbitrap technology, is both novel and unique, and no such instrument is currently available within the UK. This would be a 'first', and we have accordingly secured a substantial contribution from the applicant Universities.

Technical Summary

Many of the projects in the newly formed Liverpool Centre for Metabolomics Research involve a requirement for the sophisticated, detailed, and RAPID analysis of biological samples to identify and measure the small many hundreds or thousands of molecules (metabolites) that they contain. This is typically done using liquid chromatography followed by high-resolution mass spectrometry, including the masses of any fragments that may be produced. Classically, a major bottleneck has been the IDENTIFICATION of the metabolites represented by what may otherwise be highly reproducible retention times and MS^n mass spectra. An instrument called the Orbitrap ID-X tribrid, together with an integrated suite of software (to which we shall add), offers a real chance of proving the necessary spectra and solving this 'spectrum-to-structure' problem. It may be applied to multiple problems within the BBSRC remit, and we illustrate several.

The aim of this proposal is to secure such an instrument for an extensive User Community in Liverpool and elsewhere. The instrument, while based on established Orbitrap technology, is both novel and unique, and no such instrument is available within the UK.

Planned Impact

WHO WILL BENEFIT: Companies will benefit in a number of ways, by (i) gaining access to knowledge of methods used for the rapid analysis of biological systems using state-of-the-art XC-MS instrumentation and its attendant analytics, and e.g. (ii) knowledge of how to improve the production of various compounds, whether they are substrates, intermediary metabolites, nutraceuticals, drugs, or bioactives in any kind of products. So far as industrial biotechnology more generally is concerned, companies will benefit from knowledge of a novel strategy for rapid cell analysis.

HOW WILL THEY BENEFIT: As is our practice, all pertinent data are made available via the Web, and OA publishing has long been our norm. We also hold frequent workshops to assist dissemination of research results. We have pioneered in the Altmetrics field for digital dissemination - indeed a recent Nature article (Kwok R: Altmetrics make their mark. Nature 2013; 500:491-492) highlighted the fact that the PI's paper Hull D, Pettifer SR, Kell DB: Defrosting the digital library: bibliographic tools for the next generation web. PLoS Comput Biol 2008; 4:e1000204 was the most accessed in ANY PLoS journal, with over 53,000 accesses! (it is past 108,000 now) - the PI's paper Kell DB: Iron behaving badly: inappropriate iron chelation as a major contributor to the aetiology of vascular and other progressive inflammatory and degenerative diseases. BMC Med Genom 2009; 2:2 has over 98,000 accesses, increasing at ~50 per day).We shall work closely with University KT staff and industrial IP offices to secure intellectual property rights for any useful inventions that we discover. Having secured IP, future development work can take place, and several routes to commercialisation can be explored. For example, all pharmaceutical companies have their own relevant groups, with whom we are in contact. Finally, having secured IP, we shall, of course, seek actively to communicate our scientific findings to the wider research community through scientific meetings, scholarly publications and press releases.

THE WIDER COMMUNITY: DBK (@dbkell) and RG (@RoyGoodacre) are well known and regular tweeters (the latter also runs the @metabolomics twitter feed), and social media will provide a novel and useful means of disseminating our findings. Both are regular attenees at thee MetaboMeeting and International Metabolomics Society's conferences.

COMMUNICATIONS: We will communicate with relevant industrial partners both directly and via the meetings of relevant learned societies (we are members of several). In year three of the Project, we will organise a half-day meeting to explain our research to interested industrial scientists. However, we will also provide a video link to facilitate the participation of
those who are unable to travel to Liverpool.

Publications

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Title Ergothioneine: the new super antioxidant 
Description Scientific animation 
Type Of Art Film/Video/Animation 
Year Produced 2020 
Impact Created to inform lay as well as professional audiences. 
URL https://www.youtube.com/watch?v=SDiYMPpbUpk
 
Description NxNW: Innovation to Commercialisation of University Research (ICURe) Programme
Amount £45,000 (GBP)
Organisation Queen's University Belfast 
Sector Academic/University
Country United Kingdom
Start 06/2020 
End 10/2020
 
Description Daye 
Organisation Daye
Country United Kingdom 
Sector Private 
PI Contribution Member of Medical Advisory Board. Approached to join due to extensive research in the area of the gut microbiome.
Collaborator Contribution The company is developing and optimising intuitive, scientifically robust sexual and reproductive health products.
Impact The company have created and marketed new products.
Start Year 2019
 
Title METHODS FOR PRODUCTION OF ERGOTHIONEINE 
Description The present invention relates to microbial factories, in particular yeast factories, for production of ergothioneine. Also provided are methods for producing ergothioneine in a yeast cell, as well as useful nucleic acids, polypeptides, vectors and host cells. 
IP Reference WO2020221795 
Protection Patent granted
Year Protection Granted 2020
Licensed No
Impact The present invention relates to microbial factories, in particular yeast factories, for production of ergothioneine. Also provided are methods for producing ergothioneine in a yeast cell, as well as useful nucleic acids, polypeptides, vectors and host cells. Ergothioneine (ERG) (2-mercaptohistidine trimethylbetaine, (2S)-3-(2-Thioxo-2,3-dihydro-1 H-imidazol-4-yl)-2-(trimethylammonio)propanoate) is a naturally occurring antioxidant that can be found universally in plants and mammals; it possesses a tautomeric structure, but is mainly present in the thione form at physiological pH. Ergothioneine displays antioxidant properties, including scavenging of free radicals and of reactive oxygen species, but also chelating of divalent metal ions. Ergothioneine has been shown to reduce oxidative damage in rats and humans. So far only some bacteria and fungi have been identified as natural producers of ergothioneine. Ergothioneine was discovered in 1909 in the ergot fungus Claviceps purpurea, and its structure was determined two years later. Later, several other organisms were found to produce ergothioneine, including the filamentous fungus Neurospora crassa, the yeast Schizosaccharomyces pombe, and various actinobacteria including Mycobacterium smegmatis. Humans must obtain ergothioneine through their diet; some mushrooms and other foods contain up to 7 mg.g-1 dry weight. Because of its beneficial effects and possible involvement in preventing disease, ergothioneine is primed to take a place in the global dietary supplement market. Studies show that ergothioneine in humans is mainly accumulated in the liver, the kidneys, in erythrocytes, bone marrow, the eye lens and seminal fluid. It is transported by SLC22A4 (previously known as OCTN1), a transporter common to most animals. The high abundance of ergothioneine in the body could indicate that ergothioneine is involved in the maintenance of health or the mitigation of disease. Ergothioneine has demonstrated effects in in vivo models of several neurodegenerative diseases, in ischaemia reperfusion injury, and in a variety of other diseases. It is also reported that ergothioneine can accumulate at sites of injury through the upregulation of SLC22A4/OCTN1. Ergothioneine is only slowly metabolized and excreted in humans, again suggesting that it plays an important role in the body. Current methods for production of ergothioneine are mostly based on chemical synthesis. Such methods are not cost-effective and also have a significant impact on the environment. Therefore, methods for cost-effective and environmental-friendly production of ergothioneine are required.
 
Description Webinar 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Kell DB (2020) How drugs really get into cells: why passive bilayer diffusion is a myth. ACS webinar, 55 min + questions, freely available online at https://www.acs.org/content/acs/en/acs-webinars/drug-discovery/so-lute-carriers.html
Year(s) Of Engagement Activity 2020
URL https://www.acs.org/content/acs/en/acs-webinars/drug-discovery/so-lute-carriers.html