Engineering Microbial Utilization of Pectin-Rich Biomass for Producing Commodity Chemicals

Each year, 1 million tons of dried orange pulp and 7 million tons of dried sugar beet pulp are produced in the United States as byproducts of the citrus juice and sugar industries. Typically, these wastes are dried, pelleted and repurposed as livestock feed, adding little value to the final product. Interestingly, these and other agricultural wastes contain large fractions of pectin. Pectin, unlike woody lignocellulosic polymers, can be efficiently hydrolyzed into monomers of its backbone sugar, D-galacturonic acid. We believe that pectin-rich wastestreams can be used as raw material for microbial fermentations, yielding value-added bioproducts. Engineering a microbial strain to convert pectin hydrolysates into commodity chemicals poses a number of challenges we have tackled. First, the low pH from high galacturonic acid concentrations was addressed by the choice of Saccharomyces cerevisiae, which is naturally robust to low pH, as the production host. Second, we addressed the ability to import galacturonic acid in the presence of high glucose concentrations also present in pectin hydrolysate. We isolated a transporter that could be heterologously expressed to accomplish efficient import at both high and low pHs without inhibition by glucose. Third, production of commodity chemicals of interest such as mucic acid, a precursor for hydroxylated nylon, resulted in a cofactor imbalance. We demonstrated that a second pathway could be introduced to balance cofactor usage to dramatically improve production titers and host viability. Together, these strain modifications yielded a host that should be capable of high titer conversion of pectin-rich waste streams into commodity chemicals of value.