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

点击翻译按钮获取中文摘要

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.