(473b) Optimizing Carbon Efficiency of Fermentation By Engineering Mixotrophic Clostridium Coculture and Interspecies Interactions.

Fermentation of carbohydrates leads to loss of one third of feedstock carbon, as CO₂, fundamentally constraining the maximum product yields. Without enhancing carbon efficiency, development of an economically competitive bioprocess for renewable commodity chemicals and biofuel production is likely difficult. To address the fundamental challenge, we engineered mixotrophic Clostridium cocultures that simultaneously consume sugar substrates with H₂ and CO₂ gases and synthesize value-added chemical products. As a proof of concept, we designed and constructed a synthetic coculture consisting of solvent producing specialist C. acetobutylicum (Cac) and CO₂ fixation specialist C. ljungdahlii (Clj). The two species physically interact with each other and exchange intracellular materials such as RNA, protein, and electron. A genetically manipulated Cac-Clj coculture system enabled carbon negative isopropanol production with yields and selectivity higher than theoretically possible in conventional fermentation. In this presentation, we will discuss critical engineering and biological factors enabling the high performance of the Clostridium coculture. We employed synthetic biology, metabolic engineering, and systems biology strategies to dissect the coculture system and identify critical factors determining stability and efficiency of the syntrophic coculture. The developed system not only helps decarbonization of chemical manufacturing industry, but also identifies design principles for developing microbial cocultures capable of performing complicated industrial tasks.

Supported by the U.S. Department of Energy ARPA-E project under contract AR0001505.

N.B.W. and J.H. were supported in part by a U.S. Department of Education GAANN Fellowship under grant P200A210065.