title: Engineering improved bio-jet fuel tolerance in Escherichia coli using a transgenic library from the hydrocarbon-degrader Marinobacter aquaeolei.
abstract: BACKGROUND
  Recent metabolic engineering efforts have generated microorganisms that can produce biofuels, including bio-jet fuels, however these fuels are often toxic to cells, limiting production yields. There are natural examples of microorganisms that have evolved mechanisms for tolerating hydrocarbon-rich environments, such as those that thrive near natural oil seeps and in oil-polluted waters.
  
  
  RESULTS
  Using genomic DNA from the hydrocarbon-degrading microbe Marinobacter aquaeolei, we constructed a transgenic library that we expressed in Escherichia coli. We exposed cells to inhibitory levels of pinene, a monoterpene that can serve as a jet fuel precursor with chemical properties similar to existing tactical fuels. Using a sequential strategy with a fosmid library followed by a plasmid library, we were able to isolate a region of DNA from the M. aquaeolei genome that conferred pinene tolerance when expressed in E. coli. We determined that a single gene, yceI, was responsible for the tolerance improvements. Overexpression of this gene placed no additional burden on the host. We also tested tolerance to other monoterpenes and showed that yceI selectively improves tolerance.
  
  
  CONCLUSIONS
  The genomes of hydrocarbon-tolerant microbes represent a rich resource for tolerance engineering. Using a transgenic library, we were able to identify a single gene that improves E. coli's tolerance to the bio-jet fuel precursor pinene.
authors.affiliation: School of Engineering, University of Vermont, 33 Colchester Ave, Burlington, VT 05405 USA.
authors.forename: Timothy A
authors.initials: TA
authors.lastname: Tomko
authors.affiliation: School of Engineering, University of Vermont, 33 Colchester Ave, Burlington, VT 05405 USA.
authors.forename: Mary J
authors.initials: MJ
authors.lastname: Dunlop
keywords: Biofuel tolerance; Genomic library; Marinobacter aquaeolei; Monoterpene; Pinene; Transgenic; YceI