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Global transcriptome response of recombinant Escherichia coli to heat-shock and dual heat-shock recombinant protein induction

  • Original Paper
  • Published:
Journal of Industrial Microbiology and Biotechnology

Abstract

Recombinant Escherichia coli cultures are used to manufacture numerous therapeutic proteins and industrial enzymes, where many of these processes use elevated temperatures to induce recombinant protein production. The heat-shock response in wild-type E. coli has been well studied. In this study, the transcriptome profiles of recombinant E. coli subjected to a heat-shock and to a dual heat-shock recombinant protein induction were examined. Most classical heat-shock protein genes were identified as regulated in both conditions. The major transcriptome differences between the recombinant and reported wild-type cultures were heavily populated by hypothetical and putative genes, which indicates recombinant cultures utilize many unique genes to respond to a heat-shock. Comparison of the dual stressed culture data with literature recombinant protein induced culture data revealed numerous differences. The dual stressed response encompassed three major response patterns: induced-like, in-between, and greater than either individual stress response. Also, there were no genes that only responded to the dual stress. The most interesting difference between the dual stressed and induced cultures was the amino acid-tRNA gene levels. The amino acid-tRNA genes were elevated for the dual cultures compared to the induced cultures. Since, tRNAs facilitate protein synthesis via translation, this observed increase in amino acid-tRNA transcriptome levels, in concert with elevated heat-shock chaperones, might account for improved productivities often observed for thermo-inducible systems. Most importantly, the response of the recombinant cultures to a heat-shock was more profound than wild-type cultures, and further, the response to recombinant protein induction was not a simple additive response of the individual stresses.

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References

  1. Baneyx F, Mujacic M (2004) Recombinant protein folding and misfolding in Escherichia coli. Nat Biotechnol 22:1399–1408

    Article  CAS  Google Scholar 

  2. Blattner FR, Plunkett G, Bloch CA, Perna NT, Burland V, Riley M, ColladoVides J, Glasner JD, Rode CK, Mayhew GF, Gregor J, Davis NW, Kirkpatrick HA, Goeden MA, Rose DJ, Mau B, Shao Y (1997) The complete genome sequence of Escherichia coli K-12. Science 277:1453–1474

    Article  CAS  Google Scholar 

  3. Bonomo J, Gill R (2005) Amino acid content of recombinant proteins influences the metabolic burden response. Biotechnol Bioeng 90:116–126

    Article  CAS  Google Scholar 

  4. Chuang S-E, Blattner FR (1993) Characterization of twenty-six new heat shock genes of Escherichia coli. J Bacteriol 175:5242–5252

    CAS  Google Scholar 

  5. Chuang SE, Daniels DL, Blattner FR (1993) Global regulation of gene-expression in Escherichia coli. J Bacteriol 175:2026–2036

    CAS  Google Scholar 

  6. DeLisa MP, Li J, Rao G, Weigand WA, Bentley WE (1999) Monitoring GFP-operon fusion protein expression during high cell density cultivation of Escherichia coli using an on-line optical sensor. Biotechnol Bioeng 65:54–64

    Article  CAS  Google Scholar 

  7. Gadgil M, Kapur V, Hu WS (2005) Transcriptional response of Escherichia coli to temprature shift. Biotechnol Prog 21:689–699

    Article  CAS  Google Scholar 

  8. Gao HC, Wang Y, Liu XD, Yan TF, Wu LY, Alm E, Arkin A, Thompson DK, Zhou JZ (2004) Global transcriptome analysis of the heat shock response of shewanella oneidensis. J Bacteriol 186:7796–7803

    Article  CAS  Google Scholar 

  9. Gerdes SY, Scholle MD, Campbell JW, Balazsi G, Ravasz E, Daugherty MD, Somera AL, Kyrpides NC, Anderson I, Gelfand MS, Bhattacharya A, Kapatral V, D’Souza M, Baev MV, Grechkin Y, Mseeh F, Fonstein MY, Overbeek R, Barabasi AL, Oltvai ZN, Osterman AL (2003) Experimental determination and system level analysis of essential genes in Escherichia coli MG1655. J Bacteriol 185:5673–5684

    Article  CAS  Google Scholar 

  10. Gill RT, Delisa MP, Valdes JJ, Bentley WE (2001) Genomic analysis of high cell density recombinant Escherichia coli fermentation and “cell conditioning” for improved recombinant protein yield. Biotechnol Bioeng 72:85–95

    Article  CAS  Google Scholar 

  11. Gill RT, Valdes JJ, Bentley WE (1999) Reverse transcription-PCR differential display analysis of Escherichia coli global gene regulation in response to heat shock. Appl Environ Microbiol 65:5386–5393

    CAS  Google Scholar 

  12. Glasner JD, Liss P, Plunkett G, Darling A, Prasad T, Rusch M, Byrnes A, Gilson M, Biehl B, Blattner FR, Perna NT (2003) ASAP, a systematic annotation package for community analysis of genomes. Nucleic Acids Res 31:147–151

    Article  CAS  Google Scholar 

  13. Gross CA (1996) Function and regulation of the heat shock. In: Neidhardt FC, Curtiss R, Lin ECC, Low KB, Magasanik B, Reanikoff WS, Riley M, Schaechter M, Umbarger HE (eds) Escherichia coli and Salmonella. ASM Press, Washington DC, pp 1382–1399

    Google Scholar 

  14. Grossman AD, Taylor WE, Burton ZF, Burgess RR, Gross CA (1985) Stringent response in Escherichia coli induces expression of heat shock proteins. J Mol Biol 186:357–365

    Article  CAS  Google Scholar 

  15. Haddadin FT, Harcum SW (2005) Transcriptome profiles for high-cell-density recombinant and wild-type Escherichia coli. Biotechnol Bioeng 90:127–153

    Article  CAS  Google Scholar 

  16. Han MJ, Park SJ, Park TJ, Lee SY (2004) Roles and applications of small heat shock proteins in the production of recombinant proteins in Escherichia coli. Biotechnol Bioeng 88:426–436

    Article  CAS  Google Scholar 

  17. Harcum SW, Bentley WE (1993) Response dynamics of 26-, 34-, 39-, 54-, and 80-kDa proteases in induced cultures of recombinant Escherichia coli. Biotechnol Bioeng 42:675–685

    Article  CAS  Google Scholar 

  18. Harcum SW, Bentley WE (1999) Heat-shock and stringent responses have overlapping protease activity in Escherichia coli: Implications for heterologous protein yield. Appl Biochem Biotechnol 80:23–37

    Article  CAS  Google Scholar 

  19. Henry MD, Yancey SD, Kushner SR (1992) Role of the heat shock response in stability of mRNA in Escherichia coli K-12. J Bacteriol 174:743–748

    CAS  Google Scholar 

  20. Herendeen SL, VanBogelen RA, Neidhardt FC (1979) Levels of major proteins of Escherichia coli during growth at different temperatures. J Bacteriol 139:185–194

    CAS  Google Scholar 

  21. Hoffmann F, Rinas U (2001) Plasmid amplification in Escherichia coli after temperature up shift is impaired by induction of recombinant protein synthesis. Biotechnol Lett 23:1819–1825

    Article  CAS  Google Scholar 

  22. Hoffmann F, Weber J, Rinas U (2002) Metabolic adaptation of Escherichia coli during temperature-induced recombinant protein production: 1. Readjustment of metabolic enzyme synthesis. Biotechnol Bioeng 80:313–319

    Article  CAS  Google Scholar 

  23. Kanemori M, Mori H, Yura T (1994) Induction of heat shock proteins by abnormal proteins results from stabilization and not increased synthesis of σ32 in Escherichia coli. J Bacteriol 176:5648–5653

    CAS  Google Scholar 

  24. Kucharczyk K, Laskowska E, Taylor A (1991) Response of Escherichia coli cell membranes to induction of l cl857 prophage by heat shock. Mol Microbiol 5:2935–2945

    Article  CAS  Google Scholar 

  25. Lee SC, Olins PO (1992) Effect of overproduction of heat shock chaperones GroESL and DnaK on human procollagenase production in Escherichia coli. J Biol Chem 267:2849–2852

    CAS  Google Scholar 

  26. Lemaux PG, Herendeen SL, Bloch P, Neidhardt FC (1978) Transient rates of synthesis of individual polypeptides in E. coli following temperature shifts. Cell 13:427–434

    Article  CAS  Google Scholar 

  27. Lesley SA, Graziano J, Cho CY, Knuth MW, Klock HE (2002) Gene expression response to misfolded protein as a screen for soluble recombinant protein. Protein Eng 15:153–160

    Article  CAS  Google Scholar 

  28. Makrides SC (1996) Strategies for achieving high-level expression of genes in Escherichia coli. Microbiol Rev 60:512

    CAS  Google Scholar 

  29. Nagai H, Yuzawa H, Yura T (1991) Regulation of the heat shock response in E. coli: involvement of positive and negative cis-acting elements in translational control of σ32 synthesis. Biochimie 73:1473–1479

    Article  CAS  Google Scholar 

  30. Pilon AL, Yost P, Chase TE, Lohnas GL, Bentley WE (1996) High-level expression and efficient recovery of ubiquitin fusion proteins from Escherichia coli. Biotechnol Prog 12:331–337

    Article  CAS  Google Scholar 

  31. Richmond CS, Glasner JD, Mau R, Jin H, Blattner FJ (1999) Genome-wide expression profiling in Escherichia coli K-12. Nucleic Acids Res 27:3821–3835

    Article  CAS  Google Scholar 

  32. Riley M, Abe T, Arnaud MB, Berlyn MKB, Blattner FR, Chaudhuri RR, Glasner JD, Horiuchi T, Keseler IM, Kosuge T, Mori H, Perna NT, Plunkett G, Rudd KE, Serres MH, Thomas GH, Thomson NR, Wishart D, Wanner BL (2006) Escherichia coli K-12: a cooperatively developed annotation snapshot - 2005. Nucleic Acids Res 34:1–9

    Article  CAS  Google Scholar 

  33. Rodriguez RL, Tait RE (1983) Recombinant DNA techniques: an introduction. Benjamin/Cummings, Menlo Park, CA

    Google Scholar 

  34. Rohlin L, Oh MK, Liao JC (2002) DNA microarray for microbial biotechnology: gene expression profiles in Escherichia coli during protein overexpression. J Chin Inst Chem Eng 33:103–112

    CAS  Google Scholar 

  35. Schmidt M, Babu KR, Khanna N, Marten S, Rinas U (1999) Temperature-induced production of recombinant human insulin in high-cell density cultures of recombinant Escherichia coli. J Biotechnol 68:71–83

    Article  CAS  Google Scholar 

  36. Schmidt M, Viaplana E, Hoffmann F, Marten S, Villaverde A, Rinas U (1999) Secretion-dependent proteolysis of heterologous protein by recombinant Escherichia coli is connected to an increased activity of the energy-generating dissimilatory pathway. Biotechnol Bioeng 66:61–67

    Article  CAS  Google Scholar 

  37. Srivastava R, Peterson MS, Bentley WE (2001) Stochastic kinetic analysis of the Escherichia coli stress circuit using σ32-targeted antisense. Biotechnol Bioeng 75:120–129

    Article  CAS  Google Scholar 

  38. Storz G, Regine H-A (2000) Bacterial stress response. ASM Press, Wachington, DC

    Google Scholar 

  39. Thomas JG, Baneyx F (1996) Influence of a global deregulation of the heat-shock response on the expression of heterologous proteins in Escherichia coli. Ann N Y Acad Sci 782:478–485

    Article  CAS  Google Scholar 

  40. Thomas JG, Baneyx F (1996) Protein folding in the cytoplasm of Escherichia coli: requirements for the DnaK-DnaJ-GrpE and GroEL-GroES molecular chaperone machines. Mol Microbiol 21:1185–1196

    Article  CAS  Google Scholar 

  41. Vlazny D, Hill C (1995) A stationary-phase-dependent viability block governed byt two different polypeptides from the rhsa genetic element of Escherichia coli K-12. J Bacteriol 177:2209–2213

    CAS  Google Scholar 

  42. Wegrzyn A, Wegrzyn G, Taylor K (1996) Disassembly of the coliphage l replication complex due to heat shock induction of the groE operon. Virology 217:594–597

    Article  CAS  Google Scholar 

  43. Wild J, Walter WA, Gross CA, Altman E (1993) Accumulation of secretory protein precursors in Escherichia coli induces the heat shock response. J Bacteriol 175:3992–3997

    CAS  Google Scholar 

  44. Xu HM, Zhang GY, Ji XD, Cao L, Shu L, Hua ZC (2005) Expression of soluble, biologically active recombinant human endostatin in Escherichia coli. Protein Expres Purif 41:252–258

    Article  CAS  Google Scholar 

  45. Yabuta M, Onaimiura S, Ohsuye K (1995) Thermo-inducible expression of a recombinant fusion protein by Escherichia coli lac repressor mutants. J Biotechnol 39:67–73

    Article  CAS  Google Scholar 

  46. Yamamori T, Ito K, Nakamura Y, Yura T (1978) Transient regulation of protein synthesis in Escherichia coli upon shift-up of growth temperature. J Bacteriol 134:1133–1140

    CAS  Google Scholar 

Download references

Acknowledgment

This material is based upon work supported by the National Science Foundation under Grant No. 0303782.

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Correspondence to Sarah W. Harcum.

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Harcum, S.W., Haddadin, F.T. Global transcriptome response of recombinant Escherichia coli to heat-shock and dual heat-shock recombinant protein induction. J IND MICROBIOL BIOTECHNOL 33, 801–814 (2006). https://doi.org/10.1007/s10295-006-0122-3

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  • DOI: https://doi.org/10.1007/s10295-006-0122-3

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