Sustainable Production of Marine Cyanobacterial Products (Sus-C-Products): discovery, scale up and maintenance

There is increasing realization of the biotechnological potential of cyanobacteria as sources of new compounds and as green factories for the bioproduction of industrial feedstocks for a wide range of industries, from bioplastics, pharmaceuticals to high value compounds such as pigments. Unlike conventional microbial discovery and production platforms, cyanobacteria are light dependent, offering novel production methods as well as being an under-explored biodiscovery resource. Advances in molecular biology and in photonics render cyanobacteria much more tractable. Using combinations of specific wavelengths of light, carefully controlled culture conditions coupled with molecular discovery techniques opens up the prospect of a much-enlarged microbial discovery pipeline which may be of use to address pressing problems such as discovery of new antibiotics for use against drug-resistant infections and low carbon manufacturing. To move from discovery to product also requires the ability to scale up production quickly and easily and to ensure that the chemical activity of interest is maintained in cultures at industrial scale. Problems in i) achieving scale up andii) loss of activity in cultures are two major industrial biotech problem areas. We propose to develop a cyanobacterial pipeline for the discovery and production of novel compounds of industrial interest: coupling discovery with production systems that scale well. Compounds isolated from microorganisms and plants provide an unparalleled starting point for new compound and enzyme discovery. Over the past 3 decades more than 70% of antibiotics entering clinical trials have been based on such compounds. Marine cyanobacteria represent a particularly rich and largely untapped source of natural products, for example producing series of valuable bioactives such as dolastatin, jamaicamide, atanapeptin. Cyanobacteria also encode the assembly commercially attractive compounds such as non-proteinogenic amino acids (eg dolamethylleucine, dolaphenvaline) for which current synthetic approaches employed are very costly. Other rare motifs, such as terminal alkynes (with great potential as tools for chemical biology) are also encoded. We will use state-of- the-art combined genomics and metabolomics to mine and discover compounds from cyanobacteria, and mediate their sustainable and green fermentative production. Biosynthetic gene clusters will be selected for their ability to mediate chemically unprecedented transformations. Unusual enzyme activities will be explored both within their associated biosynthetic clusters as well as being excised and developed as tools for biotransformations.3
Rather than using traditional grind and find methods of discovery, we propose a state of the art, genome led approach: specifically we will:
1. combine genomic, metabolic and bioactivity analysis to identify biosynthetic gene clusters (BGCs). that encode enzymes with the potential to biosynthesise structural novel natural products with .medicinally useful activities .
2. use appropriate combinations of heterologous expression and promotor refactoring within the Wild Type and heterologous host (specifically tailored to each system) to unlock the production of .unusual enzymes and novel natural products .
3. natural products will be isolated purified and structurally characterized using cutting edge approached that combine detailed mass spectrometric analysis, genome analysis and 1 and 2DNMR experiments (this will provide training for the fellow in structural characterization. .
4. sustainable, fermentative production of the compounds from both the refactored WT systems and the heterologous host is key. The impact of large scale culturing on strain stability will be assessed using Xanthella's range of culturing systems that allow cultures to be grown under very controlled conditions and at scales ranging from 1l to growth in large scale PBRs at 600L.