Engineering microbial biofuel tolerance and export using efflux pumps

Mol Syst Biol. 2011 May 10;7:487. doi: 10.1038/msb.2011.21.

Abstract

Many compounds being considered as candidates for advanced biofuels are toxic to microorganisms. This introduces an undesirable trade-off when engineering metabolic pathways for biofuel production because the engineered microbes must balance production against survival. Cellular export systems, such as efflux pumps, provide a direct mechanism for reducing biofuel toxicity. To identify novel biofuel pumps, we used bioinformatics to generate a list of all efflux pumps from sequenced bacterial genomes and prioritized a subset of targets for cloning. The resulting library of 43 pumps was heterologously expressed in Escherichia coli, where we tested it against seven representative biofuels. By using a competitive growth assay, we efficiently distinguished pumps that improved survival. For two of the fuels (n-butanol and isopentanol), none of the pumps improved tolerance. For all other fuels, we identified pumps that restored growth in the presence of biofuel. We then tested a beneficial pump directly in a production strain and demonstrated that it improved biofuel yields. Our findings introduce new tools for engineering production strains and utilize the increasingly large database of sequenced genomes.

Publication types

  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • 1-Butanol* / metabolism
  • 1-Butanol* / toxicity
  • Biofuels* / toxicity
  • Computational Biology
  • Escherichia coli / genetics*
  • Escherichia coli / growth & development
  • Escherichia coli / metabolism*
  • Genetic Engineering / methods*
  • Membrane Transport Proteins / genetics*
  • Membrane Transport Proteins / metabolism
  • Metabolic Networks and Pathways
  • Microarray Analysis
  • Pentanols* / metabolism
  • Pentanols* / toxicity

Substances

  • Biofuels
  • Membrane Transport Proteins
  • Pentanols
  • 1-Butanol
  • isopentyl alcohol