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Improving olefin tolerance and production in E. coli using native and evolved AcrB.

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Engineering microbes for bioplastics production is challenging due to toxic compounds. This study shows the AcrAB-TolC efflux pump enhances tolerance to olefins like styrene and 1-hexene in E. coli, with directed evolution improving this further.

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directed evolutionhost engineeringolefin productionsolvent tolerance

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Area of Science:

  • Microbial Engineering
  • Biotechnology
  • Biopolymer Production

Background:

  • Microbial production of polymer precursors like styrene and alpha-olefins is hindered by compound toxicity.
  • The AcrAB-TolC efflux pump in Escherichia coli plays a role in tolerating these toxic compounds.

Purpose of the Study:

  • To investigate the role of the AcrAB-TolC efflux pump in enhancing tolerance to styrene and alpha-olefins (1-hexene, 1-octene, 1-nonene) in E. coli.
  • To improve tolerance to 1-hexene using directed evolution targeting the AcrB component of the efflux pump.

Main Methods:

  • Evaluated the role of the AcrAB-TolC efflux pump in E. coli tolerance to styrene, 1-hexene, 1-octene, and 1-nonene.
  • Employed directed evolution to optimize the AcrB protein for enhanced 1-hexene tolerance, focusing on mutations in the substrate-binding domain.

Main Results:

  • AcrAB-TolC contributes to E. coli tolerance towards all tested olefin compounds.
  • Styrene and 1-hexene exhibited high toxicity to E. coli.
  • Directed evolution identified mutations in AcrB (A279T, Q584R, F617L, L822P, F927S, F1033Y) that significantly improved 1-hexene tolerance, with synergistic effects observed in combined mutations.

Conclusions:

  • Efflux pumps are crucial for host engineering strategies aimed at producing olefins for the polymer industry.
  • Directed evolution is a viable strategy to enhance efflux pump-mediated tolerance, potentially increasing bioplastic precursor production.