Engineering a Probiotic Bacterium for Utilization of Pyrolytic Sugars

Fermentable sugar obtained from lignocellulosic material exhibits substantial potential as a renewable feedstock. Upon pyrolysis, this biomass produces a major product called levoglucosan (LG), which is not fermented by typical microbes such as Escherichia coli. Previous reports suggest that engineering microbes with the levoglucosan kinase (Lgk) enzyme can enable the use of this sugar, directing carbon flux toward growth and energy metabolism. In this study, we have engineered the probiotic strain E.coli Nissle 1917 (EcN) for LG utilization with recombinant expression of Lgk enzyme from Lipomyces starkeyi. We demonstrated inducible and constitutive expression of Lgk in EcN, and increasing protein expression enabled use of LG up to 10% (w/v) as the sole carbon source in minimal media. This engineered strain also showed a competitive growth advantage when co-cultured with organisms lacking Lgk. Our current investigation involves constructing a metabolic pathway for the production of anti-inflammatory molecules like butyrate, secreting anti-bacterial peptides, and identifying transporters that will enable more efficient catabolism of pyrolytic sugars.