Award details

17-ERACoBioTech Thermophilic bacteria and archaeal chassis for extremolyte production -HotSolute

Reference	BB/R02166X/1
Principal Investigator / Supervisor	Professor Jennifer Littlechild
Co-Investigators / Co-Supervisors	Professor Nicholas Harmer, Dr Mikhail Isupov
Institution	University of Exeter
Department	Biosciences
Funding type	Research
Value (£)	412,012
Status	Completed
Type	Research Grant
Start date	01/03/2018
End date	31/03/2022
Duration	49 months

Abstract

The proposed project will establish thermophilic in vitro enzyme cascades as well as two new chassis, the thermophilic bacterium Thermus thermophilus (Tth, 65-75degrees C, pH 7.0) and the thermoacidophilic archaeon Sulfolobus acidocaldarius (Saci, 75-80 degrees C, pH 2-4), as new thermophilic, bacterial and archaeal platforms for the production of novel high added-value products, extremolytes. The development of the newly designed 'cell factories' will be used for the production of three extremolytes, cyclic 2,3 di-phosphoglycerate(cDPG), di-myo-1,1'-inositol-phosphate (DIP) and mannosylglycerate (MG). These extremolytes (with few exceptions for MG) are exclusively found in hyperthermophiles, and have not been produced in a mesophilic host to date. The project will use these existing natural biological cell based systems with a synthetic biology approach. New biological so called parts and devices will be employed in order to establish and optimize two new thermophilic 'cell factories' for the production of the novel extremolytes with medical and personal care applications. Both organisms, Saci and Tth are easy to grow (minimal or complex media, aerobic growth). Importantly advanced genetic tools have been established for both Tth and Saci that will allow for the insertion of new modules using a synthetic biology approach. For enzyme cascade and strain design, construction, optimization and product recovery, a model-based systems biology and synthetic biology approach will be employed including state of the art genetics, biochemistry, transcriptomics, proteomics, modelling, data management and life cycle assessment. During the project a bioinformatic approach will also be used to search for new enzymes from metagenomes and newly sequenced hyper-thermophilic genomes. To provide small quantities of extremolytes to the industrial partners (SME and Evonik) for testing early in the project by isolation from the natural producer strain.

Summary

Thermophilic organisms are composed of both bacterial and archaeal species. The enzymes isolated from these species and from other extreme habitats are more robust to high temperature, organic solvents and to breakdown from other proteolytic enzymes. They often have unique substrate specificities and originate from novel metabolic pathways. Thermophiles as well as their stable enzymes ('thermozymes') are receiving increased attention for biotechnological applications. The proposed project will establish thermophilic in vitro enzyme cascades as well as two new chassis, the thermophilic bacterium Thermus thermophilus (Tth) and the thermoacidophilic archaeon Sulfolobus acidocaldarius (Saci) as new thermophilic, bacterial and archaeal platforms for the production of novel high added-value products called 'extremolytes'. Extremolytes are small molecule compatible solutes found naturally in the cells of thermophilic species that accumulate in the cell in response to multiple environmental stresses and help to stabilize cellular components (including proteins and membranes). Extremolytes offer an amazing so far unexploited potential for industrial applications including food, health, consumer care and cosmetics. However, their production in common mesophilic organisms such as fungi and Escherichia coli is currently hampered by the hyperthermophilic origin of the respective metabolic pathways which require a thermophilic cell factory. The development of the newly designed 'cell factories' will be used for the production of three extremolytes, cyclic 2,3 di-phosphoglycerate (cDPG), di-myo-1,1'-inositol-phosphate (DIP) and mannosylglycerate (MG). These extremolytes (with few exceptions for MG) are exclusively found in hyperthermophiles, and have not been produced in a mesophilic host to date. The extremolyte biosynthetic pathways have been identified and many of the enzymes involved have been characterized. Within the project in addition to these well established enzymes,new candidates will be provided by searching for new synthetic enzymes in metagenome newly isolated strains from (hyper)thermophilic habitats. All three extremolytes are derived in a few steps from central glycolytic intermediates and are absent in Saci and only MG has been reported in Tth. The establishment of thermophilic in vitro enzyme cascades as well as in vivo enzyme platforms will be used for extremolyte production. Both organisms, Saci and Tth are easy to grow (minimal or complex media, aerobic growth). Many other thermophilic organisms require anaerobic or specialised conditions to achieve successful growth in the laboratory or in an industrial setting. Importantly advanced genetic tools have been established for both Tth and Saci that will allow the insertion of new modules into the cells using a synthetic biology approach. For enzyme cascade and strain design, construction, optimization and product recovery a model-based systems biology and synthetic biology approach will be employed including state of the art genetics, biochemistry, transcriptomics, proteomics, modelling, data management and life cycle assessment.

Impact Summary

The impact This project will develop the application of thermophilic micro-organisms (both bacterial and archaeal) as cell factories for the production of new industrially interesting molecules. Thermophilic enzyme cascades will be developed in order to produce three high added-value products extremolytes, which have applications in medical and skin care products. The platforms once established can be used for production of other important products using a synthetic biology approach of enzyme cascades for both in vivo and in vitro systems and drive new applications of industrial biotechnology. Natural producer strains will be used to produce extremolytes for initial testing The public visibility and dissemination events will make people increasingly aware of the importance and possibilities that research and innovation in industrial biotechnology can offer for them. Increased knowledge in this area will be transferred from academia to industry and will enable training of experienced personnel in this multidisciplinary area of sustainable bio-processing. This project directly contributes to the aims of this ERA-CoBioTech call, to transform the global economy from a dependence on fossil raw materials to a sustainable bio-based economy. The development of this approach will enable future developments and innovations in response to societal and industrial needs within the bioeconomy. In the longer term, it will further propagate the exploitation of the hot bacterial and archaeal chassis for the integration of other new synthetic modules that have direct applications of industrial biotechnology. The combination of new thermophilic features (e.g. pathways, enzymes and regulatory circuits) of bacterial, eukaroytic and archaeal origin offer new exciting possibilities for future applications in industrial biotechnology. The production of the three new high-value compounds, extremolytes, cDPG, DIP and MG represents great innovative potential for the cosmetics and health care industries. The large scale production of extremolytes will trigger technological innovation by new patents for extremolyte employment and increase the established market for other extremolytes (e.g. ectoine, hydroxyectoine) in the cosmetic and pharmaceutical industries. These compounds are known to stabilize proteins, cell membranes and produce a lung surfactant monolayer. They are also used in anti-allergic creams as well as moisturizing nasal sprays, eye drops, inhalation solutions, derma products and as anti-aging and sun protection cosmetics. In this respect they will improve the overall health of the general public.

Committee	Not funded via Committee
Research Topics	Industrial Biotechnology, Microbiology, Synthetic Biology
Research Priority	X – Research Priority information not available
Research Initiative	ERA-Net Cofund on Biotechnologies (ERACoBioTech) [2017]
Funding Scheme	X – not Funded via a specific Funding Scheme

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