Genome Organisation for Optimising Synthetic Secondary Metabolism

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This project will research genome organisation and in particular how changing the location and arrangement of metabolic enzyme genes within a yeast genome can alter the amount of the metabolite they produce, in this case the antibiotic penicillin. Antibiotics are just one class of complex chemicals that the diverse array of organisms on Earth has naturally evolved to produce. The production of chemicals by life is known as metabolism and the more complex high-value chemicals that specialist cells produce (e.g. in plants) are called secondary metabolites and these include most therapeutic molecules known today. Production of secondary metabolites in cells requires specific enzymes which are encoded by genes usually under strict control (regulation). As our understanding of biology improves through many fundamental research breakthroughs, scientists are now looking to re-engineer secondary metabolism to produce valuable compounds in cellular systems that are easy to work with. Microbes like brewer's yeast are perfect as they are easy to culture and so could cheaply produce high yields of valuable compounds from renewable resources like sugar.The most promising way to perform this 'metabolic engineering' is to use what is known as a synthetic biology approach, where genes and their controllers are treated as modular components with well-defined behaviours and then combined in a rational design-based manner. So far the synthetic biology approach to metabolic engineering has been successful in producing compounds useful as anti-malarials, cosmetics and biofuels by taking genes for enzymes found in plants and exotic microbes and combining these inside industrially-used microbes such as yeast. Crucial for achieving high yields of production in these microbes is fine-control over the precise levels of enzymes in each cell.One method of tuning enzyme levels that is currently unexplored by scientists is how the genes for these enzymes are physically arranged within a cell's genome. It is already known that gene location and orientation within a genome plays an important role in gene expression in all forms of life. Recently, it has also been established that in cells that naturally perform secondary metabolism, such as plant cells, the location of genes that make up a pathway is often tightly conserved, usually in occurring in gene 'clusters' in areas known as 'sub-telomeric regions'. Clearly, if nature and evolution are correct, then the location of where pathway genes are added to a genome must affect the enzyme levels and therefore the pathway output.This project seeks to test the hypothesis that modifying pathway gene location within a genome can result in improved yields of a high-value secondary metabolite, in this case penicillin. The genes encoding the penicillin pathway will be added to the genome of a lab yeast strain and cells will be selected that produce the greatest amounts of penicillin. The amount produced will then be monitored in a series of experiments where the pathway genes are systematically rearranged around five different places in the genome. This will give valuable information on how the arrangement of genes in the genome affects the pathway. Finally, the pathway genes will be placed in an engineered lab strain specifically designed to shuffle parts of its genome when under an evolutionary pressure. The strain will be grown to compete against bacteria and in doing will automatically rearrange pathway genes to produce the most penicillin. This project will therefore provide an important new synthetic biology approach to metabolic engineering, and also uncover valuable new information on the fundamental science of genomes and genome evolution.