Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Perspective
  • Published:

Stress-tolerant non-conventional microbes enable next-generation chemical biosynthesis

Nature Chemical Biology volume 16, pages 113–121 (2020)Cite this article

Subjects

Abstract

Microbial chemical production is a rapidly growing industry, with much of the growth fueled by advances in synthetic biology. New approaches have enabled rapid strain engineering for the production of various compounds; however, translation to industry is often problematic because native phenotypes of model hosts prevent the design of new low-cost bioprocesses. Here, we argue for a new approach that leverages the native stress-tolerant phenotypes of non-conventional microbes that directly address design challenges from the outset. Growth at high temperature, high salt and solvent concentrations, and low pH can enable cost savings by reducing the energy required for product separation, bioreactor cooling, and maintaining sterile conditions. These phenotypes have the added benefit of allowing for the use of low-cost sugar and water resources. Non-conventional hosts are needed because these phenotypes are polygenic and thus far have proven difficult to recapitulate in the common hosts Escherichia coli and Saccharomyces cerevisiae.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Fig. 1: Membrane chemistries of stress-tolerant microbes.
Fig. 2: Metabolism and transport of the osmolyte ectoine.
Fig. 3: High solvent tolerance in Pseudomonas putida through effective efflux pumping.
Fig. 4: Progress of synthetic biology tool development.

Similar content being viewed by others

References

  1. Carlson, R. Estimating the biotech sector’s contribution to the US economy. Nat. Biotechnol. 34, 247–255 (2016). This Perspective article provides a detailed analysis of the US industrial biotechnology sector.

    Article  CAS  PubMed  Google Scholar 

  2. Jinek, M. et al. A programmable dual-RNA-guided DNA endonuclease in adaptive bacterial immunity. Science 337, 816–821 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Caruthers, M. H. A brief review of DNA and RNA chemical synthesis. Biochem. Soc. Trans. 39, 575–580 (2011).

    Article  CAS  PubMed  Google Scholar 

  4. Chao, R., Mishra, S., Si, T. & Zhao, H. Engineering biological systems using automated biofoundries. Metab. Eng. 42, 98–108 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Hong, K. K. & Nielsen, J. Metabolic engineering of Saccharomyces cerevisiae: a key cell factory platform for future biorefineries. Cell. Mol. Life Sci. 69, 2671–2690 (2012).

    Article  CAS  PubMed  Google Scholar 

  6. Pontrelli, S. et al. Escherichia coli as a host for metabolic engineering. Metab. Eng. 50, 16–46 (2018).

    Article  CAS  PubMed  Google Scholar 

  7. Shapouri, H. & Gallagher, P. USDA’s 2002 Ethanol Cost-of-Production Survey. Agricultural Economics Report Number 841 (US Department of Agriculture, Office of Energy Policy and New Uses, 2005).

  8. Blanch, H.W. & Clark, D.S. Biochemical Engineering, xii (M. Dekker, 1996).

  9. Lam, F. H., Ghaderi, A., Fink, G. R. & Stephanopoulos, G. Biofuels. Engineering alcohol tolerance in yeast. Science 346, 71–75 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Rich, J. O., Leathers, T. D., Bischoff, K. M., Anderson, A. M. & Nunnally, M. S. Biofilm formation and ethanol inhibition by bacterial contaminants of biofuel fermentation. Bioresour. Technol. 196, 347–354 (2015).

    Article  CAS  PubMed  Google Scholar 

  11. Löbs, A. K., Engel, R., Schwartz, C., Flores, A. & Wheeldon, I. CRISPR–Cas9-enabled genetic disruptions for understanding ethanol and ethyl acetate biosynthesis in Kluyveromyces marxianus. Biotechnol. Biofuels 10, 164 (2017).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  12. Shui, W. et al. Understanding the mechanism of thermotolerance distinct from feat shock response through proteomic analysis of industrial strains of Saccharomyces cerevisiae. Mol. Cell. Proteom. 14, 1885–1897 (2015).

    Article  CAS  Google Scholar 

  13. Richter, K., Haslbeck, M. & Buchner, J. The heat shock response: life on the verge of death. Mol. Cell 40, 253–266 (2010).

    Article  CAS  PubMed  Google Scholar 

  14. Phipps, B. M. et al. Structure of a molecular chaperone from a thermophilic Archaebacterium. Nature 361, 475–477 (1993).

    Article  CAS  Google Scholar 

  15. Parsell, D. A. & Lindquist, S. The function of heat-shock proteins in stress tolerance: degradation and reactivation of damaged proteins. Annu. Rev. Genet. 27, 437–496 (1993).

    Article  CAS  PubMed  Google Scholar 

  16. Strassburg, K., Walther, D., Takahashi, H., Kanaya, S. & Kopka, J. Dynamic transcriptional and metabolic responses in yeast adapting to temperature stress. OMICS 14, 249–259 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Balchin, D., Hayer-Hartl, M. & Hartl, F. U. In vivo aspects of protein folding and quality control. Science 353, aac4354 (2016).

    Article  PubMed  CAS  Google Scholar 

  18. Verghese, J., Abrams, J., Wang, Y. & Morano, K. A. Biology of the heat shock response and protein chaperones: budding yeast (Saccharomyces cerevisiae) as a model system. Microbiol. Mol. Biol. Rev. 76, 115–158 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Gasch, A. P. et al. Genomic expression programs in the response of yeast cells to environmental changes. Mol. Biol. Cell 11, 4241–4257 (2000).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Rodrigues, J. L. & Rodrigues, L. R. Potential applications of the Escherichia coli heat shock response in synthetic biology. Trends Biotechnol. 36, 186–198 (2018).

    Article  CAS  PubMed  Google Scholar 

  21. Lewin, A., Wentzel, A. & Valla, S. Metagenomics of microbial life in extreme temperature environments. Curr. Opin. Biotechnol. 24, 516–525 (2013).

    Article  CAS  PubMed  Google Scholar 

  22. Jacquemet, A., Barbeau, J., Lemiègre, L. & Benvegnu, T. Archaeal tetraether bipolar lipids: structures, functions and applications. Biochimie 91, 711–717 (2009).

    Article  CAS  PubMed  Google Scholar 

  23. Li, P. et al. The transcription factors Hsf1 and Msn2 of thermotolerant Kluyveromyces marxianus promote cell growth and ethanol fermentation of Saccharomyces cerevisiae at high temperatures. Biotechnol. Biofuels 10, 289 (2017).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  24. Seong, Y. J. et al. Physiological and metabolomic analysis of Issatchenkia orientalis MTY1 with multiple tolerance for cellulosic bioethanol production. Biotechnol. J. 12, 1700110 (2017).

    Article  CAS  Google Scholar 

  25. Blaby, I. K. et al. Experimental evolution of a facultative thermophile from a mesophilic ancestor. Appl. Environ. Microbiol. 78, 144–155 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Rudolph, B., Gebendorfer, K. M., Buchner, J. & Winter, J. Evolution of Escherichia coli for growth at high temperatures. J. Biol. Chem. 285, 19029–19034 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Caspeta, L. et al. Biofuels. Altered sterol composition renders yeast thermotolerant. Science 346, 75–78 (2014).

    Article  CAS  PubMed  Google Scholar 

  28. Shi, D. J., Wang, C. L. & Wang, K. M. Genome shuffling to improve thermotolerance, ethanol tolerance and ethanol productivity of Saccharomyces cerevisiae. J. Ind. Microbiol. Biotechnol. 36, 139–147 (2009).

    Article  CAS  PubMed  Google Scholar 

  29. Bhandiwad, A. et al. Metabolic engineering of Thermoanaerobacterium saccharolyticum for n-butanol production. Metab. Eng. 21, 17–25 (2014).

    Article  CAS  PubMed  Google Scholar 

  30. Lin, P. P. et al. Consolidated bioprocessing of cellulose to isobutanol using Clostridium thermocellum. Metab. Eng. 31, 44–52 (2015).

    Article  CAS  PubMed  Google Scholar 

  31. Cripps, R. E. et al. Metabolic engineering of Geobacillus thermoglucosidasius for high yield ethanol production. Metab. Eng. 11, 398–408 (2009).

    Article  CAS  PubMed  Google Scholar 

  32. Yue, H. T. et al. A seawater-based open and continuous process for polyhydroxyalkanoates production by recombinant Halomonas campaniensis LS21 grown in mixed substrates. Biotechnol. Biofuels 7, 108 (2014). This paper shows the power of stress-tolerant microbes. Engineered halophilic H. campaniensis was used in non-aseptic bioreactors to produce high titers of polyhydroxybutyrate from mixed substrate feedstocks.

    Article  CAS  Google Scholar 

  33. Voronovsky, A. Y., Rohulya, O. V., Abbas, C. A. & Sibirny, A. A. Development of strains of the thermotolerant yeast Hansenula polymorpha capable of alcoholic fermentation of starch and xylan. Metab. Eng. 11, 234–242 (2009).

    Article  CAS  PubMed  Google Scholar 

  34. Löbs, A. K., Lin, J. L., Cook, M. & Wheeldon, I. High throughput, colorimetric screening of microbial ester biosynthesis reveals high ethyl acetate production from Kluyveromyces marxianus on C5, C6, and C12 carbon sources. Biotechnol. J. 11, 1274–1281 (2016).

    Article  PubMed  CAS  Google Scholar 

  35. Xiao, H., Shao, Z., Jiang, Y., Dole, S. & Zhao, H. Exploiting Issatchenkia orientalis SD108 for succinic acid production. Microb. Cell Fact. 13, 121 (2014).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  36. Löbs, A. K., Schwartz, C., Thorwall, S. & Wheeldon, I. Highly multiplexed CRISPRi repression of respiratory functions enhances mitochondrial localized ethyl acetate biosynthesis in Kluyveromyces marxianus. ACS Synth. Biol. 7, 2647–2655 (2018). This paper demonstrates the power of CRISPR technologies for rapidly engineering non-conventional hosts. Here, multiplexed CRISPR interference (CRISPRi) was used to screen various transcriptional programs to optimize a native biosynthetic pathway in the thermotolerant yeast K. marxianus.

    Article  PubMed  CAS  Google Scholar 

  37. Abdel-Banat, B. M., Hoshida, H., Ano, A., Nonklang, S. & Akada, R. High-temperature fermentation: how can processes for ethanol production at high temperatures become superior to the traditional process using mesophilic yeast? Appl. Microbiol. Biotechnol. 85, 861–867 (2010).

    Article  CAS  PubMed  Google Scholar 

  38. Cernak, P. et al. Engineering Kluyveromyces marxianus as a robust synthetic biology platform host. MBio 9, e01410–e01418 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Cheon, Y. et al. A biosynthetic pathway for hexanoic acid production in Kluyveromyces marxianus. J. Biotechnol. 182-183, 30–36 (2014).

    Article  CAS  PubMed  Google Scholar 

  40. Dmytruk, K., Kurylenko, O., Ruchala, J., Ishchuk, O. & Sibirny, A. in Yeast Diversity in Human Welfare (eds Satyanarayana, T. & Kunze, G.) 257–282 (Springer, 2017).

  41. Lenton, S., Walsh, D. L., Rhys, N. H., Soper, A. K. & Dougan, L. Structural evidence for solvent-stabilisation by aspartic acid as a mechanism for halophilic protein stability in high salt concentrations. Phys. Chem. Chem. Phys. 18, 18054–18062 (2016).

    Article  CAS  PubMed  Google Scholar 

  42. Oren, A. Microbial life at high salt concentrations: phylogenetic and metabolic diversity. Saline Syst. 4, 2 (2008).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  43. Padan, E., Venturi, M., Gerchman, Y. & Dover, N. Na+/H+ antiporters. Biochim. Biophys. Acta 1505, 144–157 (2001).

    Article  CAS  PubMed  Google Scholar 

  44. Purvis, J. E., Yomano, L. P. & Ingram, L. O. Enhanced trehalose production improves growth of Escherichia coli under osmotic stress. Appl. Environ. Microbiol. 71, 3761–3769 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Yang, L. et al. A primary sodium pump gene of the moderate halophile Halobacillus dabanensis exhibits secondary antiporter properties. Biochem. Biophys. Res. Commun. 346, 612–617 (2006).

    Article  CAS  PubMed  Google Scholar 

  46. Ekberg, J. et al. Adaptive evolution of the lager brewing yeast Saccharomyces pastorianus for improved growth under hyperosmotic conditions and its influence on fermentation performance. FEMS Yeast Res. 13, 335–349 (2013).

    Article  CAS  PubMed  Google Scholar 

  47. Schweikhard, E. S., Kuhlmann, S. I., Kunte, H. J., Grammann, K. & Ziegler, C. M. Structure and function of the universal stress protein TeaD and its role in regulating the ectoine transporter TeaABC of Halomonas elongata DSM 2581(T). Biochemistry 49, 2194–2204 (2010).

    Article  CAS  PubMed  Google Scholar 

  48. Kunte, H., Lentzen, G. & Galinski, E. Industrial production of the cell protectant ectoine: protection mechanisms, processes, and products. Curr. Biotechnol. 3, 10–25 (2014).

    Article  CAS  Google Scholar 

  49. Chen, X., Yu, L., Qiao, G. & Chen, G. Q. Reprogramming Halomonas for industrial production of chemicals. J. Ind. Microbiol. Biotechnol. 45, 545–554 (2018).

    Article  CAS  PubMed  Google Scholar 

  50. Qin, Q. et al. CRISPR/Cas9 editing genome of extremophile Halomonas spp. Metab. Eng. 47, 219–229 (2018).

    Article  CAS  PubMed  Google Scholar 

  51. Chen, R. et al. Optimization of the extraction and purification of the compatible solute ectoine from Halomonas elongate in the laboratory experiment of a commercial production project. World J. Microbiol. Biotechnol. 33, 116 (2017).

    Article  PubMed  CAS  Google Scholar 

  52. Zaky, A. S., Greetham, D., Tucker, G. A. & Du, C. The establishment of a marine focused biorefinery for bioethanol production using seawater and a novel marine yeast strain. Sci. Rep. 8, 12127 (2018).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  53. Yuan, W. J., Zhao, X. Q., Ge, X. M. & Bai, F. W. Ethanol fermentation with Kluyveromyces marxianus from Jerusalem artichoke grown in salina and irrigated with a mixture of seawater and freshwater. J. Appl. Microbiol. 105, 2076–2083 (2008).

    Article  CAS  PubMed  Google Scholar 

  54. Ramos, J. L., Duque, E., Godoy, P. & Segura, A. Efflux pumps involved in toluene tolerance in Pseudomonas putida DOT-T1E. J. Bacteriol. 180, 3323–3329 (1998).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  55. Rühl, J., Schmid, A. & Blank, L. M. Selected Pseudomonas putida strains able to grow in the presence of high butanol concentrations. Appl. Environ. Microbiol. 75, 4653–4656 (2009).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  56. Ramos, J. L. et al. Mechanisms for solvent tolerance in bacteria. J. Biol. Chem. 272, 3887–3890 (1997).

    Article  CAS  PubMed  Google Scholar 

  57. Rojas, A. et al. Three efflux pumps are required to provide efficient tolerance to toluene in Pseudomonas putida DOT-T1E. J. Bacteriol. 183, 3967–3973 (2001).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  58. Terán, W. et al. Complexity in efflux pump control: cross-regulation by the paralogues TtgV and TtgT. Mol. Microbiol. 66, 1416–1428 (2007).

    PubMed  Google Scholar 

  59. Segura, A. et al. Proteomic analysis reveals the participation of energy- and stress-related proteins in the response of Pseudomonas putida DOT-T1E to toluene. J. Bacteriol. 187, 5937–5945 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  60. Dunlop, M. J. et al. Engineering microbial biofuel tolerance and export using efflux pumps. Mol. Syst. Biol. 7, 487 (2011).

    Article  PubMed  PubMed Central  Google Scholar 

  61. Tan, Z., Yoon, J. M., Nielsen, D. R., Shanks, J. V. & Jarboe, L. R. Membrane engineering via trans unsaturated fatty acids production improves Escherichia coli robustness and production of biorenewables. Metab. Eng. 35, 105–113 (2016).

    Article  CAS  PubMed  Google Scholar 

  62. Zingaro, K. A. & Terry Papoutsakis, E. GroESL overexpression imparts Escherichia coli tolerance to i-, n-, and 2-butanol, 1,2,4-butanetriol and ethanol with complex and unpredictable patterns. Metab. Eng. 15, 196–205 (2013).

    Article  CAS  PubMed  Google Scholar 

  63. Cook, T. B. et al. Genetic tools for reliable gene expression and recombineering in Pseudomonas putida. J. Ind. Microbiol. Biotechnol. 45, 517–527 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  64. Gong, T. et al. Metabolic engineering of Pseudomonas putida KT2440 for complete mineralization of methyl parathion and γ-hexachlorocyclohexane. ACS Synth. Biol. 5, 434–442 (2016).

    Article  CAS  PubMed  Google Scholar 

  65. Nikel, P. I. & de Lorenzo, V. Pseudomonas putida as a functional chassis for industrial biocatalysis: from native biochemistry to trans-metabolism. Metab. Eng. 50, 142–155 (2018).

    Article  CAS  PubMed  Google Scholar 

  66. Bormann, S. et al. Engineering Clostridium acetobutylicum for production of kerosene and diesel blendstock precursors. Metab. Eng. 25, 124–130 (2014).

    Article  CAS  PubMed  Google Scholar 

  67. Anbarasan, P. et al. Integration of chemical catalysis with extractive fermentation to produce fuels. Nature 491, 235–239 (2012).

    Article  CAS  PubMed  Google Scholar 

  68. Zhang, J., Wu, C. D., Du, G. C. & Chen, J. Enhanced acid tolerance in Lactobacillus casei by adaptive evolution and compared stress response during acid stress. Biotechnol. Bioprocess Eng. 17, 283–289 (2012).

    Article  CAS  Google Scholar 

  69. Zhou, L. et al. Improvement of d-lactate productivity in recombinant Escherichia coli by coupling production with growth. Biotechnol. Lett. 34, 1123–1130 (2012).

    Article  CAS  PubMed  Google Scholar 

  70. Lee, J. Y., Kang, C. D., Lee, S. H., Park, Y. K. & Cho, K. M. Engineering cellular redox balance in Saccharomyces cerevisiae for improved production of l-lactic acid. Biotechnol. Bioeng. 112, 751–758 (2015).

    Article  CAS  PubMed  Google Scholar 

  71. Yan, D. et al. Construction of reductive pathway in Saccharomyces cerevisiae for effective succinic acid fermentation at low pH value. Bioresour. Technol. 156, 232–239 (2014).

    Article  CAS  PubMed  Google Scholar 

  72. Kwon, Y. D., Kim, S., Lee, S. Y. & Kim, P. Long-term continuous adaptation of Escherichia coli to high succinate stress and transcriptome analysis of the tolerant strain. J. Biosci. Bioeng. 111, 26–30 (2011).

    Article  CAS  PubMed  Google Scholar 

  73. Wright, J. et al. Batch and continuous culture-based selection strategies for acetic acid tolerance in xylose-fermenting Saccharomyces cerevisiae. FEMS Yeast Res. 11, 299–306 (2011).

    Article  CAS  PubMed  Google Scholar 

  74. Sandoval, N. R., Mills, T. Y., Zhang, M. & Gill, R. T. Elucidating acetate tolerance in E. coli using a genome-wide approach. Metab. Eng. 13, 214–224 (2011).

    Article  CAS  PubMed  Google Scholar 

  75. Suominen, P. et al. Genetically modified yeast of the species Issatchenkia Orientalis and closely relates species, and fermentation processes using same (Cargill Inc., 2012).

  76. Valdés, J. et al. Acidithiobacillus ferrooxidans metabolism: from genome sequence to industrial applications. BMC Genomics 9, 597 (2008).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  77. Kernan, T. et al. Engineering the iron-oxidizing chemolithoautotroph Acidithiobacillus ferrooxidans for biochemical production. Biotechnol. Bioeng. 113, 189–197 (2016).

    Article  CAS  PubMed  Google Scholar 

  78. Inaba, Y., Banerjee, I., Kernan, T. & Banta, S. Transposase-mediated chromosomal integration of exogenous genes in Acidithiobacillus ferrooxidans. Appl. Environ. Microbiol. 84, e01381–18 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  79. Maezato, Y., Johnson, T., McCarthy, S., Dana, K. & Blum, P. Metal resistance and lithoautotrophy in the extreme thermoacidophile Metallosphaera sedula. J. Bacteriol. 194, 6856–6863 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  80. Auernik, K. S., Maezato, Y., Blum, P. H. & Kelly, R. M. The genome sequence of the metal-mobilizing, extremely thermoacidophilic archaeon Metallosphaera sedula provides insights into bioleaching-associated metabolism. Appl. Environ. Microbiol. 74, 682–692 (2008).

    Article  CAS  PubMed  Google Scholar 

  81. Zeldes, B. M. et al. Extremely thermophilic microorganisms as metabolic engineering platforms for production of fuels and industrial chemicals. Front. Microbiol. 6, 1209 (2015).

    Article  PubMed  PubMed Central  Google Scholar 

  82. Feist, A. M. et al. A genome-scale metabolic reconstruction for Escherichia coli K-12 MG1655 that accounts for 1260 ORFs and thermodynamic information. Mol. Syst. Biol. 3, 121 (2007).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  83. Förster, J., Famili, I., Fu, P., Palsson, B. O. & Nielsen, J. Genome-scale reconstruction of the Saccharomyces cerevisiae metabolic network. Genome Res. 13, 244–253 (2003).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  84. Wang, H. H. et al. Programming cells by multiplex genome engineering and accelerated evolution. Nature 460, 894–898 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  85. Warner, J. R., Reeder, P. J., Karimpour-Fard, A., Woodruff, L. B. & Gill, R. T. Rapid profiling of a microbial genome using mixtures of barcoded oligonucleotides. Nat. Biotechnol. 28, 856–862 (2010).

    Article  CAS  PubMed  Google Scholar 

  86. Lee, M. E., DeLoache, W. C., Cervantes, B. & Dueber, J. E. A highly characterized yeast toolkit for modular, multipart assembly. ACS Synth. Biol. 4, 975–986 (2015).

    Article  CAS  PubMed  Google Scholar 

  87. Xu, P., Vansiri, A., Bhan, N. & Koffas, M. A. ePathBrick: a synthetic biology platform for engineering metabolic pathways in E. coli. ACS Synth. Biol. 1, 256–266 (2012).

    Article  CAS  PubMed  Google Scholar 

  88. Löbs, A. K., Schwartz, C. & Wheeldon, I. Genome and metabolic engineering in non-conventional yeasts: current advances and applications. Synth. Syst. Biotechnol. 2, 198–207 (2017).

    Article  PubMed  PubMed Central  Google Scholar 

  89. Schwartz, C., Frogue, K., Ramesh, A., Misa, J. & Wheeldon, I. CRISPRi repression of nonhomologous end-joining for enhanced genome engineering via homologous recombination in Yarrowia lipolytica. Biotechnol. Bioeng. 114, 2896–2906 (2017).

    Article  CAS  PubMed  Google Scholar 

  90. Cao, M., Gao, M., Ploessl, D., Song, C. & Shao, Z. CRISPR-mediated genome editing and gene repression in Scheffersomyces stipitis. Biotechnol. J. 13, e1700598 (2018).

    Article  PubMed  CAS  Google Scholar 

  91. Schwartz, C., Shabbir-Hussain, M., Frogue, K., Blenner, M. & Wheeldon, I. Standardized markerless gene integration for pathway engineering in Yarrowia lipolytica. ACS Synth. Biol. 6, 402–409 (2017).

    Article  CAS  PubMed  Google Scholar 

  92. Schwartz, C. et al. Validating genome-wide CRISPR-Cas9 function improves screening in the oleaginous yeast Yarrowia lipolytica. Metab. Eng. 55, 102–110 (2019).

    Article  CAS  PubMed  Google Scholar 

  93. Schwartz, C. M., Hussain, M. S., Blenner, M. & Wheeldon, I. Synthetic RNA polymerase III promoters facilitate high-efficiency CRISPR-Cas9-mediated genome editing in Yarrowia lipolytica. ACS Synth. Biol. 5, 356–359 (2016).

    Article  CAS  PubMed  Google Scholar 

  94. Cao, M. et al. Centromeric DNA facilitates nonconventional yeast genetic engineering. ACS Synth. Biol. 6, 1545–1553 (2017).

    Article  CAS  PubMed  Google Scholar 

  95. Lewis, N. E., Nagarajan, H. & Palsson, B. O. Constraining the metabolic genotype-phenotype relationship using a phylogeny of in silico methods. Nat. Rev. Microbiol. 10, 291–305 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  96. Schwalen, C. J., Hudson, G. A., Kille, B. & Mitchell, D. A. Bioinformatic expansion and discovery of thiopeptide antibiotics. J. Am. Chem. Soc. 140, 9494–9501 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  97. Brophy, J. A. N. et al. Engineered integrative and conjugative elements for efficient and inducible DNA transfer to undomesticated bacteria. Nat. Microbiol. 3, 1043–1053 (2018). This work engineers the integrative and conjugative elements of Bacillus subtilis to create a strain technology called XPORT that can be used to rapidly engineer novel bacterial isolates.

    Article  CAS  PubMed  Google Scholar 

  98. Schwartz, C., Curtis, N., Löbs, A. K. & Wheeldon, I. Multiplexed CRISPR activation of cryptic sugar metabolism enables Yarrowia lipolytica growth on cellobiose. Biotechnol. J. 13, e1700584 (2018).

    Article  PubMed  CAS  Google Scholar 

  99. Boundy-Mills, K. L. et al. Yeast culture collections in the twenty-first century: new opportunities and challenges. Yeast 33, 243–260 (2016).

    Article  CAS  PubMed  Google Scholar 

  100. Solomon, K. V. et al. Early-branching gut fungi possess a large, comprehensive array of biomass-degrading enzymes. Science 351, 1192–1195 (2016). This work uses ‘omics’-level analyses and biochemical assays to discover new biotechnology-relevant enzymes, including cellulases for the breakdown of lignocellulosic biomass in new isolates of anaerobic gut fungi.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

This material is based upon work supported by the US Department of Energy, Office of Science, Office of Biological and Environmental Research, Genomic Science Program under Award Number DE-SC0019093, Air Force Office of Scientific Research award FA9550-17-1-0270, Army Research Office MURI award W911NF1410263, and National Science Foundation award NSF-CBET 1706545 for funding.

Author information

Authors and Affiliations

  1. Department of Chemical and Environmental Engineering, University of California Riverside, Riverside, CA, USA

    Sarah Thorwall, Cory Schwartz & Ian Wheeldon

  2. Department of Bioengineering, University of California Riverside, Riverside, CA, USA

    Justin W. Chartron

  3. Center for Industrial Biotechnology, Bourns College of Engineering, University of California Riverside, Riverside, CA, USA

    Ian Wheeldon

Authors
  1. Sarah Thorwall
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Cory Schwartz
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Justin W. Chartron
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Ian Wheeldon
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Ian Wheeldon.

Ethics declarations

Competing interests

The authors declare no competing interests.

Additional information

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Thorwall, S., Schwartz, C., Chartron, J.W. et al. Stress-tolerant non-conventional microbes enable next-generation chemical biosynthesis. Nat Chem Biol 16, 113–121 (2020). https://doi.org/10.1038/s41589-019-0452-x

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/s41589-019-0452-x

This article is cited by

Search

Advanced search

Quick links

Nature Briefing: Translational Research

Sign up for the Nature Briefing: Translational Research newsletter — top stories in biotechnology, drug discovery and pharma.

Get what matters in translational research, free to your inbox weekly. Sign up for Nature Briefing: Translational Research