Stephan Lane¹, Shuyan Zhang^2,3, Cristopher Rao^2,3, and Yong-Su Jin^1,2

¹Department of Food Science and Human Nutrition, University of Illinois at Urbana-Champaign

² Carl R. Woese Institute for Genomic Biology, University of Illinois at Urbana-Champaign

³Department of Chemical and Biomolecular Engineering, University of Illinois at Urbana-Champaign
Lipids and organic acids are often used in the food industry as important flavor compounds and stabilizers.  However, production of these compounds may rely on fishing, farming, and other ecologically-damaging processes.  Alternatively, chemical synthesis can offer a bypass for these ecological concerns, however consumers often find artificial ingredients unappealing.  Fortunately, the growing field of metabolic engineering opens new directions for efficient and natural production of these industrially-useful compounds.  To enable cost-effective production of lipids and organic acids, we sought to enable biosynthesis of these useful ingredients from the most abundant organic polymer on the planet:  cellulose.  Here we describe a process for microbial digestion of the dimeric component of cellulose by engineering the yeast Yarrowia lipolytica to express a cellodextrin transporter and β-glucosidase.  By controlling growth conditions, we are able to guide the engineered yeast’s metabolism to produce either lipids or organic acids.  We apply this technology in an enhanced process of simultaneous saccharification and fermentation, wherein cellulose is directly converted to useful products.  As a proof of concept, we first demonstrate that engineered Yarrowia lipolytica is capable of digesting the dimeric form of cellulose, cellobiose.  In addition to being capable of sustaining cell growth, the dimeric sugar can be metabolically converted into both lipids and citric acid.  Surprisingly, we find that this conversion takes place at rates equal to—and in some cases even surpassing—sugars naturally consumed by the species.  Ultimately, we demonstrate direct biotransformation of cellulose to citric acid by culturing our engineered yeast with crystalline cellulose supplemented with endoglucanase. We conclude that digestion of cellulose by genetically engineered microorganisms reveals untapped potential for bioconversion of this extremely abundant polymer into products and ingredients important to the food industry.