P105: A novel pathway for propionyl-CoA production in E. coli

The synthesis of propionyl-CoA is required for several metabolic pathways that produce value-added products, including polyketide synthesis and the pentanol biosynthetic pathway that was recently described in our lab (Tseng and Prather. 2012. PNAS, 109:17925).  Additionally, propionyl-CoA can be reduced to 1-propanol in a two-step reaction catalyzed by the bi-functional alcohol/aldehyde dehydrogenase, AdhE.  Propanol serves as an industrial solvent and can be dehydrated to propylene, a precursor to the plastic polypropylene.  Propionyl-CoA, however, is not naturally produced by E. coli.  Here we demonstrate a novel pathway that reduces glycerol to propionyl-CoA, which can be subsequently reduced to 1-propanol.  This pathway is initiated by the radical-enzyme glycerol dehydratase, which forms 3-hydroxypropionaldehyde.  Next, a series of two CoA-mediated reductions then produce propionyl-CoA.  By decoupling expression of the oxygen-sensitive radical-activator from the rest of the pathway genes, flux through glycerol dehydratase was significantly increased.  Furthermore, by placing the activator under control of a native anoxic E. coli promoter and by constitutively expressing the remainder of the pathway genes, an autonomous pathway was established which required no exogenous chemical inducers for expression.  To further increase flux through the recombinant pathway, multiplex genome engineering with co-selection was utilized to alter the expression of several native E. coli genes.  The engineered strains possessed up to ten-fold improvement in pathway flux.