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Title: Guanidine Riboswitch-Regulated Efflux Transporters Protect Bacteria against Ionic Liquid Toxicity

Abstract

Plant cell walls contain a renewable, nearly limitless supply of sugar that could be used to support microbial production of commodity chemicals and biofuels. Imidazolium ionic liquid (IIL) solvents are among the best reagents for gaining access to the sugars in this otherwise recalcitrant biomass. However, the sugars from IIL-treated biomass are inevitably contaminated with residual IILs that inhibit growth in bacteria and yeast, blocking biochemical production by these organisms. IIL toxicity is, therefore, a critical roadblock in many industrial biosynthetic pathways. Although several IIL-tolerant (IILT) bacterial and yeast isolates have been identified in nature, few genetic mechanisms have been identified. In this study, we identified two IILT Bacillus isolates as well as a spontaneous IILT Escherichia coli lab strain that are tolerant to high levels of two widely used IILs. We demonstrate that all three IILT strains contain one or more pumps of the small multidrug resistance (SMR) family, and two of these strains contain mutations that affect an adjacent regulatory guanidine riboswitch. Furthermore, we show that the regulation of E. coli sugE by the guanidine II riboswitch can be exploited to promote IIL tolerance by the simple addition of guanidine to the medium. Our results demonstrate the criticalmore » role that transporter genes play in IIL tolerance in their native bacterial hosts. The study presented here is another step in engineering IIL tolerance into industrial strains toward overcoming this key gap in biofuels and industrial biochemical production processes.« less

Authors:
 [1];  [2]; ORCiD logo [3];  [4];  [4]; ORCiD logo [1]
  1. Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)
  2. Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Sandia National Lab. (SNL-CA), Livermore, CA (United States)
  3. Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)
  4. Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States)
Publication Date:
Research Org.:
Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)
Sponsoring Org.:
USDOE National Nuclear Security Administration (NNSA)
OSTI Identifier:
1544473
Report Number(s):
LLNL-JRNL-767148
Journal ID: ISSN 0021-9193; 957444
Grant/Contract Number:  
AC52-07NA27344
Resource Type:
Accepted Manuscript
Journal Name:
Journal of Bacteriology
Additional Journal Information:
Journal Volume: 201; Journal Issue: 13; Journal ID: ISSN 0021-9193
Publisher:
American Society for Microbiology
Country of Publication:
United States
Language:
English
Subject:
09 BIOMASS FUELS

Citation Formats

Higgins, Douglas A., Gladden, John M., Kimbrel, Jeff A., Simmons, Blake A., Singer, Steven W., and Thelen, Michael P. Guanidine Riboswitch-Regulated Efflux Transporters Protect Bacteria against Ionic Liquid Toxicity. United States: N. p., 2019. Web. doi:10.1128/JB.00069-19.
Higgins, Douglas A., Gladden, John M., Kimbrel, Jeff A., Simmons, Blake A., Singer, Steven W., & Thelen, Michael P. Guanidine Riboswitch-Regulated Efflux Transporters Protect Bacteria against Ionic Liquid Toxicity. United States. doi:10.1128/JB.00069-19.
Higgins, Douglas A., Gladden, John M., Kimbrel, Jeff A., Simmons, Blake A., Singer, Steven W., and Thelen, Michael P. Mon . "Guanidine Riboswitch-Regulated Efflux Transporters Protect Bacteria against Ionic Liquid Toxicity". United States. doi:10.1128/JB.00069-19. https://www.osti.gov/servlets/purl/1544473.
@article{osti_1544473,
title = {Guanidine Riboswitch-Regulated Efflux Transporters Protect Bacteria against Ionic Liquid Toxicity},
author = {Higgins, Douglas A. and Gladden, John M. and Kimbrel, Jeff A. and Simmons, Blake A. and Singer, Steven W. and Thelen, Michael P.},
abstractNote = {Plant cell walls contain a renewable, nearly limitless supply of sugar that could be used to support microbial production of commodity chemicals and biofuels. Imidazolium ionic liquid (IIL) solvents are among the best reagents for gaining access to the sugars in this otherwise recalcitrant biomass. However, the sugars from IIL-treated biomass are inevitably contaminated with residual IILs that inhibit growth in bacteria and yeast, blocking biochemical production by these organisms. IIL toxicity is, therefore, a critical roadblock in many industrial biosynthetic pathways. Although several IIL-tolerant (IILT) bacterial and yeast isolates have been identified in nature, few genetic mechanisms have been identified. In this study, we identified two IILT Bacillus isolates as well as a spontaneous IILT Escherichia coli lab strain that are tolerant to high levels of two widely used IILs. We demonstrate that all three IILT strains contain one or more pumps of the small multidrug resistance (SMR) family, and two of these strains contain mutations that affect an adjacent regulatory guanidine riboswitch. Furthermore, we show that the regulation of E. coli sugE by the guanidine II riboswitch can be exploited to promote IIL tolerance by the simple addition of guanidine to the medium. Our results demonstrate the critical role that transporter genes play in IIL tolerance in their native bacterial hosts. The study presented here is another step in engineering IIL tolerance into industrial strains toward overcoming this key gap in biofuels and industrial biochemical production processes.},
doi = {10.1128/JB.00069-19},
journal = {Journal of Bacteriology},
number = 13,
volume = 201,
place = {United States},
year = {2019},
month = {6}
}

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