Keynote Talk: Engineering Complex Metabolic Pathways for Secondary Metabolite Production

While initial metabolic engineering research was predominantly oriented towards optimizing existing metabolic networks and concentrated predominantly on reaction networks close to central metabolism, the drive to generate higher value-added products has led to far more advanced strategies being initiated. Yields are often quite low and new challenges are faced as the modified pathways often involve intrinsically low activities for some or many of the reactions as well as requiring the optimisation of quite long reaction networks. While a few laboratories have the high throughput robotics to test multiple pathway constructions, many cannot meet this investment and need a strategy for rapid optimisation to optimize pathway flux. We have developed a modular approach which enables multi-level adjustment with relatively few constructs to maximise pathway functionality. While this involves tunable promoters for gene expression, the high yields we have achieved for apocarotenoids have also needed some specific protein engineering strategies. The approach and the results obtained will be presented as regards our efforts with apocarotenoid and terpenoid pathways and how some of these strains have been grown in biphasic fermentation to attain nature-enhanced and economically viable levels of flavour and fragrance ingredients using E. coli as a platform organism.