Elimination of Non-Productive Fermentation Products Boosts Cellulosic Ethanol Production in Consolidated Bioprocessing


Clostridium thermocellum has the natural ability to convert cellulose to ethanol, making it a promising candidate for consolidated bioprocessing (CBP) of cellulosic biomass to biofuels. In addition to ethanol, however, C. thermocellum produces a number of unwanted fermentation products such as organic acids and gaseous hydrogen, which divert energy and carbon from the desired fermentation product, ethanol. Researchers at the Department of Energy’s BioEnergy Science Center sought to eliminate these non-target fermentation products in order to increase ethanol yields. In doing so, they created C. thermocellum strain AG553 by deleting genes involved in the production of acetate, formate, lactate, and hydrogen gas. Strain AG553 showed a two- to three-fold increase in ethanol yield relative to the wild type on all substrates tested. When grown in a defined medium with 5 g/L of soluble disaccharide cellobiose as the carbon source, the mutant strain produced greater than two-fold more ethanol than the wild type strain. It exceeded 70% of theoretical ethanol yield with no appreciable amounts of other fermentation products detected and H2 production reduced five-fold. Wild type C. thermocellum will naturally acidify a non-buffered medium during fer­mentation by production of organic acids and limit ethanol production by limiting growth. The elimination of organic acid production suggested that strain AG553 might be capable of growth under higher substrate loadings in the absence of pH control. The maximum titer of wild type C. thermocellum was only 14.1 mM ethanol on 10 g/L Avicel. For strain AG553, final ethanol titer peaked at 73.4 mM in on 20 g/L Avicel, at which point the pH decreased to a level that does not allow growth of C. thermocellum, likely due to carbon dioxide accumulation. With the elimination of the non-target fermentation metabolic pathways, AG553 is the best ethanol-yielding CBP strain to date. It will serve as a platform strain for further metabolic engineering for the bioconversion of lignocellulosic biomass into advanced biofuels other than ethanol.


Papanek, B., R. Biswas, T. Rydzak, and A. M. Guss. 2015. “Elimination of Metabolic Pathways to All Traditional Fermentation Products Increases Ethanol Yields in Clostridium thermocellum,”Metabolic Engineering, DOI: 10.10/16/j.ymben.2015.09.002.