Increased microbial butanol tolerance by exogenous membrane insertion molecules

Abstract

Butanol is an ideal biofuel, although poor titers lead to high re- covery costs by distillation. Fluidization of microbial mem- branes by butanol is one of the major factors limiting titers in butanol-producing bioprocesses. Starting with the hypothesis that certain membrane insertion molecules would stabilize the lipid bilayer in the presence of butanol, we applied a combina- tion of in vivo and in vitro techniques within an in silico frame- work to describe a new approach to achieve solvent tolerance in bacteria. Single-molecule tracking of a model supported bi- layer showed that COE1-5C, a five-ringed oligo-polyphenylene- vinylene conjugated oligoelectrolyte (COE), reduced the diffu- sion rate of phospholipids in a microbially derived lipid bilayer to a greater extent than three-ringed and four-ringed COEs. Furthermore, COE1-5C treatment increased the specific growth rate of E. coli K12 relative to a control at inhibitory butanol concentrations. Consequently, to confer butanol tolerance to microbes by exogenous means is complementary to genetic modification of strains in industrial bioprocesses, extends the physiological range of microbes to match favorable bioprocess conditions, and is amenable with complex and undefined mi- crobial consortia for biobutanol production. Molecular dynam- ics simulations indicated that the p-conjugated aromatic back- bone of COE1-5C likely acts as a hydrophobic tether for glycer- ophospholipid acyl chains by enhancing bilayer integrity in the presence of high butanol concentrations, which thereby coun- ters membrane fluidization. COE1-5C-mitigated E. coli K12 membrane depolarization by butanol is consistent with the hy- pothesis that improved growth rates in the presence of buta- nol are a consequence of improved bilayer stability.