Biofuels: Anaerobes do it better!

Concerns about the stability of petroleum supply and sustainability, as well as a desire to decrease carbon emissions have resulted in an increased interest in biofuels development. A common approach involves the design of novel pathways for improved biofuels production, which are ultimately expressed in aerobic organisms, such as E. coli. The efficiency of alcohol production in these organisms is historically low when compared to native alcohol producers, likely due to the fact that these organisms are not well adapted to solventogenesis. Two such examples are the introduction of keto-isovalerate dehydrogenase (KIVD) and an alcohol dehydrogenase from yeast (Adh6) to produce isobutanol and/or 3-methylpentanol, and the introduction of betaketothiolase (BktB) from Cupriavidus necator and an aldehyde/alcohol dehydrogenase from C. acetobutylicum to produce hexanol. Both of these synthetic pathways were expressed in E. coli. Our current project involves developing a shuttle vector for expression of these and similar pathways in clostridia species that are native alcohol producers, such as C. acetobutylicum and C. beijerinckii. By operating these pathways in organisms that natively produce solvents, we hope to show an improvement over previous attempts in non-native producers of biofuels.

We have cloned several parts for constructing this shuttle vector, including the replicon and macrolide-lincosamide-streptogramin B resistance (mlsR) marker from the plasmid pIKM1, known to function in C. acetobutylicum, as well as several promoters from C. acetobutylicum. The shuttle vector will be assembled using the iGEM standard pSB1C3 plasmid backbone. We intend to express alcohol dehydrogenase genes and/or genes for one or more of the synthetic biofuels production pathways in C. acetobutylicum to demonstrate improved production of alcohols.