Metabolic engineering of Rhodopseudomonas palustris for the obligate reduction of n-butyrate to n-butanol

Devin F. R. Doud, Eric C. Holmes, Hanno Richter, Bastian Molitor, Georg Jander, Largus T. Angenent

Rhodopseudomonas palustris is a versatile microbe that encounters an innate redox imbalance while growing photoheterotrophically with reduced substrates. The resulting excess in reducing equivalents, together with ATP from photosynthesis, could be utilized to drive a wide range of bioconversions. The objective of this study was to genetically modify R. palustris to provide a pathway to reduce n-butyrate into n-butanol for maintaining redox balance. Here, we constructed and expressed a plasmid-based pathway for n-butanol production from Clostridium acetobutylicum ATCC 824 in R. palustris. We maintained the environmental conditions in such a way that this pathway functioned as the obligate route to re-oxidize excess reducing equivalents, resulting in an innate selection pressure. The engineered strain of R. palustris grew under otherwise restrictive redox conditions and achieved concentrations of 1.5 mM n-butanol at a production rate of 0.03 g L(-1) day(-1) and a selectivity (i.e., products compared to the consumed substrate) of close to 40%. Since the theoretical maximum selectivity is 45%, the engineered strain converted close to its maximum selectivity. The innate redox imbalance of R. palustris can be used to drive the reduction of n-butyrate into n-butanol after expression of a plasmid-based enzyme from a butanol-producing Clostridium strain.