Applied Microbiology and Biotechnology

, Volume 99, Issue 5, pp 2267–2275 | Cite as

Development of a novel uric-acid-responsive regulatory system in Escherichia coli

  • Chaoning Liang
  • Dandan Xiong
  • Yi Zhang
  • Shanshan Mu
  • Shuang-Yan Tang
Applied genetics and molecular biotechnology


A novel uric-acid-responsive regulatory system was developed in Escherichia coli by adapting the HucR-related regulatory elements from Deinococcus radiodurans into E. coli. The induction performance of this system was compared to the performance of both the pBAD and pET systems. Our novel regulatory system was induced in a dose-dependent manner in the presence of uric acid and exhibited low basal expression in its absence. The system was characterized by a wide dynamic range of induction, being compatible with various E. coli strains and not requiring genomic modifications of the bacterial host. E. coli DH5α and DH10B were the most suitable host strains for optimal performance of this system. In conclusion, we developed a regulatory system with potential for applications in both recombinant protein expression and metabolic optimization.


Regulatory protein Uric acid Dynamic range Induction fold Escherichia coli 



The authors would like to thank Mr. Ye Mao for his technical assistance in this research. This work was supported by the Ministry of Science and Technology of China Grant 2013CB734003, the National Natural Science Foundation of China Grant 21172095, and the Key Research Program of the Chinese Academy of Sciences Grant KSZD-EW-Z-015-2.

Supplementary material

253_2014_6290_MOESM1_ESM.pdf (88 kb)
ESM 1 (PDF 88 kb)


  1. Aaron N, Milton W (1957) Enzyme induction as an all-or-none phenomenon. Proc Natl Acad Sci U S A 43:553–566CrossRefGoogle Scholar
  2. Amann E, Brosius J, Ptashne M (1983) Vectors bearing a hybrid trp-lac promoter useful for regulated expression of cloned genes in Escherichia coli. Gene 25:167–178CrossRefPubMedGoogle Scholar
  3. Amann E, Ochs B, Abel K-J (1988) Tightly regulated tac promoter vectors useful for the expression of unfused and fused proteins in Escherichia coli. Gene 69:301–315CrossRefPubMedGoogle Scholar
  4. Baneyx F (1999) Recombinant protein expression in Escherichia coli. Curr Opin Biotechnol 10:411–421CrossRefPubMedGoogle Scholar
  5. Barrick D, Villanueba K, Childs J, Kalil R, Schneider TD, Lawrence CE, Gold L, Stormo GD (1994) Quantitative analysis of ribosome binding sites in E. coli. Nucleic Acids Res 22:1287–1295CrossRefPubMedCentralPubMedGoogle Scholar
  6. Battista JR (1997) Against all odds: the survival strategies of Deinococcus radiodurans. Annu Rev Microbiol 51:203–224CrossRefPubMedGoogle Scholar
  7. Blazeck J, Alper HS (2013) Promoter engineering: recent advances in controlling transcription at the most fundamental level. Biotechnol J 8:46–58CrossRefPubMedGoogle Scholar
  8. Carrier TA, Keasling JD (1999) Investigating autocatalytic gene expression systems through mechanistic modeling. J Theor Biol 201:25–36CrossRefPubMedGoogle Scholar
  9. Choi YJ, Morel L, Le François T, Bourque D, Bourget L, Groleau D, Massie B, Míguez CB (2010) Novel, versatile, and tightly regulated expression system for Escherichia coli strains. Appl Environ Microbiol 76:5058–5066CrossRefPubMedCentralPubMedGoogle Scholar
  10. Cox MM, Battista JR (2005) Deinococcus radiodurans - the consummate survivor. Nat Rev Microbiol 3:882–892CrossRefPubMedGoogle Scholar
  11. Datsenko KA, Wanner BL (2000) One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products. Proc Natl Acad Sci U S A 97:6640–6645CrossRefPubMedCentralPubMedGoogle Scholar
  12. Figge J, Wright C, Collins CJ, Roberts TM, Livingston DM (1988) Stringent regulation of stably integrated chloramphenicol acetyl transferase genes by E. coli lac repressor in monkey cells. Cell 52:713–722CrossRefPubMedGoogle Scholar
  13. Guan L, Liu Q, Li C, Zhang Y (2013) Development of a Fur-dependent and tightly regulated expression system in Escherichia coli for toxic protein synthesis. BMC Biotechnol 13:25–33CrossRefPubMedCentralPubMedGoogle Scholar
  14. Guido NJ, Wang X, Adalsteinsson D, McMillen D, Hasty J, Cantor CR, Elston TC, Collins JJ (2006) A bottom-up approach to gene regulation. Nature 439:856–860CrossRefPubMedGoogle Scholar
  15. Gupta JC, Jaisani M, Pandey G, Mukherjee KJ (1999) Enhancing recombinant protein yields in Escherichia coli using the T7 system under the control of heat inducible λPL promoter. J Biotechnol 68:125–134CrossRefPubMedGoogle Scholar
  16. Guzman LM, Belin D, Carson MJ, Beckwith J (1995) Tight regulation, modulation, and high-level expression by vectors containing the arabinose PBAD promoter. J Bacteriol 177:4121–4130PubMedCentralPubMedGoogle Scholar
  17. Hannig G, Makrides SC (1998) Strategies for optimizing heterologous protein expression in Escherichia coli. Trends Biotechnol 16:54–60CrossRefPubMedGoogle Scholar
  18. Harley C, Reynolds RP (1987) Analysis of E. coli promoter sequences. Nucleic Acids Res 5:2343–2361CrossRefGoogle Scholar
  19. Hooper DC, Spitsin S, Kean RB, Champion JM, Dickson GM, Chaudhry I, Koprowski H (1998) Uric acid, a natural scavenger of peroxynitrite, in experimental allergic encephalomyelitis and multiple sclerosis. Proc Natl Acad Sci U S A 95:675–680CrossRefPubMedCentralPubMedGoogle Scholar
  20. Karin H, Ruhdal JP (1998) Artificial promoter libraries for selected organisms and promoters derived from such libraries. Patent No. WO1998007846Google Scholar
  21. Keasling JD (1999) Gene-expression tools for the metabolic engineering of bacteria. Trends Biotechnol 17:452–460CrossRefPubMedGoogle Scholar
  22. Khlebnikov A, Keasling JD (2002) Effect of lacY expression on homogeneity of induction from the Ptac and Ptrc promoters by natural and synthetic inducers. Biotechnol Prog 18:672–674CrossRefPubMedGoogle Scholar
  23. Lee SK, Keasling JD (2005) A propionate-inducible expression system for enteric bacteria. Appl Environ Microbiol 71:6856–6862CrossRefPubMedCentralPubMedGoogle Scholar
  24. Lee T, Krupa R, Zhang F, Hajimorad M, Holtz W, Prasad N, Lee S, Keasling J (2011) BglBrick vectors and datasheets: a synthetic biology platform for gene expression. J Biol Eng 5:1–14CrossRefGoogle Scholar
  25. Lutz R, Bujard H (1997) Independent and tight regulation of transcriptional units in Escherichia coli via the LacR/O, the TetR/O and AraC/I1-I2 regulatory elements. Nucleic Acids Res 25:1203–1210CrossRefPubMedCentralPubMedGoogle Scholar
  26. Maeda H, Fujita N, Ishihama A (2000) Competition among seven Escherichia coli σ subunits: relative binding affinities to the core RNA polymerase. Nucleic Acids Res 28:3497–3503CrossRefPubMedCentralPubMedGoogle Scholar
  27. Nocadello S, Swennen E (2012) The new pLAI (lux regulon based auto-inducible) expression system for recombinant protein production in Escherichia coli. Microb Cell Fact 11:3–12CrossRefPubMedCentralPubMedGoogle Scholar
  28. Noh K-H, Son J-W, Kim H-J, Oh D-K (2009) Ginsenoside compound K production from ginseng root extract by a thermostable β-glycosidase from Sulfolobus solfataricus. Biosci Biotechnol Biochem 73:316–321CrossRefPubMedGoogle Scholar
  29. Papakostas K, Frillingos S (2012) Substrate selectivity of YgfU, a uric acid transporter from Escherichia coli. J Biol Chem 287:15684–15695CrossRefPubMedCentralPubMedGoogle Scholar
  30. Peti W, Page R (2007) Strategies to maximize heterologous protein expression in Escherichia coli with minimal cost. Protein Expr Purif 51:1–10CrossRefPubMedGoogle Scholar
  31. Samuelson J (2011) Recent developments in difficult protein expression: a guide to E. coli strains, promoters, and relevant host mutations. In: Evans JTC, Xu M-Q (eds) Heterologous gene expression in E. coli. Humana Press, New York, pp 195–209CrossRefGoogle Scholar
  32. Siegele DA, Hu JC (1997) Gene expression from plasmids containing the araBAD promoter at subsaturating inducer concentrations represents mixed populations. Proc Natl Acad Sci U S A 94:8168–8172Google Scholar
  33. Stark MJR (1987) Multicopy expression vectors carrying the lac represser gene for regulated high-level expression of genes in Escherichia coli. Gene 51:255–267CrossRefPubMedGoogle Scholar
  34. Studier FW, Moffatt BA (1986) Use of bacteriophage T7 RNA polymerase to direct selective high-level expression of cloned genes. J Mol Biol 189:113–130CrossRefPubMedGoogle Scholar
  35. Tabor S (2001) Expression using the T7 RNA polymerase/promoter system. In: Ausubel FA, Brent R, Kingston RE, Moore DD, Seidman JG, Smith JA, Struhl K (eds) Current protocols in molecular biology. Wiley, New York, pp 16.2.1–16.2.11Google Scholar
  36. Wagner S, Klepsch MM, Schlegel S, Appel A, Draheim R, Tarry M, Högbom M, van Wijk KJ, Slotboom DJ, Persson JO, de Gier J-W (2008) Tuning Escherichia coli for membrane protein overexpression. Proc Natl Acad Sci U S A 105:14371–14376CrossRefPubMedCentralPubMedGoogle Scholar
  37. Wilkinson SP, Grove A (2004) HucR, a novel uric acid-responsive member of the MarR family of transcriptional regulators from Deinococcus radiodurans. J Biol Chem 279:51442–51450CrossRefPubMedGoogle Scholar
  38. Wilkinson SP, Grove A (2005) Negative cooperativity of uric acid binding to the transcriptional regulator HucR from Deinococcus radiodurans. J Mol Biol 350:617–630CrossRefPubMedGoogle Scholar
  39. Zhang F, Carothers JM, Keasling JD (2012) Design of a dynamic sensor-regulator system for production of chemicals and fuels derived from fatty acids. Nat Biotechnol 30:354–359CrossRefPubMedGoogle Scholar
  40. Zheng L, Baumann U, Reymond J-L (2004) An efficient one-step site-directed and site-saturation mutagenesis protocol. Nucleic Acids Res 32:115–122CrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2014

Authors and Affiliations

  • Chaoning Liang
    • 1
  • Dandan Xiong
    • 1
    • 2
  • Yi Zhang
    • 1
    • 2
  • Shanshan Mu
    • 1
    • 2
  • Shuang-Yan Tang
    • 1
  1. 1.CAS Key Laboratory of Microbial Physiological and Metabolic Engineering, Institute of MicrobiologyChinese Academy of SciencesBeijingChina
  2. 2.University of Chinese Academy of SciencesBeijingChina

Personalised recommendations