In Silico Analysis Of Synthetic Multispecies Biofilms For Cellobiose-To-Isobutanol Conversion Reveals Design Principles For Stable And Productive Communities

Efficient, large-scale conversion of plant-derived feedstocks to commodity chemicals remains a substantial technological challenge with enormous potential for societal benefits. Most research efforts have focused on metabolic engineering of model organisms for planktonic cell cultures in well-mixed suspension bioreactors. We utilized in silico metabolic modeling to explore the potential benefits of an alternative design strategy based on combining bacterial strains with complementary metabolic functions within high density, multispecies biofilms. We simulated four alternative system designs, each of which consisted of an anaerobic cellulolytic bacterium which degraded cellobiose to glucose, an aerobic Escherichia coli strain engineered for glucose-to-isobutanol conversion, and an aerobic or anaerobic byproduct consumer for metabolizing growth-inhibiting organic acids such as acetate secreted by the other two strains. Our simulations predicted dramatically different cellobiose-toisobutanol conversion capabilities depending on the metabolic compatibility of the three bacteria. Important design considerations included glucose competition between the cellulolytic and isobutanol-producing bacteria, O-2 competition between the isobutanol-producing and byproduct-consuming bacteria, organic acid matching between the cellulolytic and byproduct-consuming bacteria and the degree of metabolic redundancy between the community members. We believe that these design principles will be widely applicable to synthetic biofilm communities engineered to perform other bioconversion tasks.