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Tracing Metabolite Footsteps of Escherichia coli Along the Time Course of Recombinant Protein Expression by Two-Dimensional NMR Spectroscopy

  • Chae, Young Kee ;
  • Kim, Seol Hyun ;
  • Ellinger, James J. ;
  • Markley, John L.
  • Received : 2012.08.14
  • Accepted : 2012.09.13
  • Published : 2012.12.20

Abstract

The recombinant expression of proteins has been the method of choice to meet the demands from proteomics and structural genomics studies. Despite its successful production of many heterologous proteins, Escherichia coli failed to produce many other proteins in their native forms. This may be related to the fact that the stresses resulting from the overproduction interfere with cellular processes. To better understand the physiological change during the overproduction phase, we profiled the metabolites along the time course of the recombinant protein expression. We identified 32 metabolites collected from different time points in the protein production phase. The stress induced by protein production can be characterized by (A) the increased usage of aspartic acid, choline, glycerol, and N-acetyllysine; and (B) the accumulation of adenosine, alanine, oxidized glutathione, glycine, N-acetylputrescine, and uracil. We envision that this work can be used to create a strategy for the production of usable proteins in large quantities.

Keywords

Metabolite profiling;Recombinant protein;Overexpression;NMR

References

  1. Baneyx, F. Curr. Opin. Biotech. 1999, 10, 411. https://doi.org/10.1016/S0958-1669(99)00003-8
  2. Oganesyan, N.; Ankoudinova, I.; Kim, S. H.; Kim, R. Protein Expr. Purif. 2007, 52, 280. https://doi.org/10.1016/j.pep.2006.09.015
  3. Tegel, H.; Steen, J.; Konrad, A.; Nikdin, H.; Pettersson, K.; Stenvall,M.; Tourle, S.; Wrethagen, U.; Xu, L.; Yderland, L.; Uhlen, M.; Hober, S.; Ottosson, J. Biotechnol. J. 2009, 4, 51. https://doi.org/10.1002/biot.200800183
  4. Prinz, W. A.; Aslund, F.; Holmgren, A.; Beckwith, J. J. Biol. Chem. 1997, 272, 15661. https://doi.org/10.1074/jbc.272.25.15661
  5. Guzman, L. M.; Belin, D.; Carson, M. J.; Beckwith, J. J. Bacteriol. 1995, 177, 4121.
  6. Dyson, M. R.; Shadbolt, S. P.; Vincent, K. J.; Perera, R. L.; McCafferty, J. BMC biotechnology 2004, 4, 32. https://doi.org/10.1186/1472-6750-4-32
  7. Niiranen, L.; Espelid, S.; Karlsen, C. R.; Mustonen, M.; Paulsen, S. M.; Heikinheimo, P.; Willassen, N. P. Protein Expr. Purif. 2007, 52, 210. https://doi.org/10.1016/j.pep.2006.09.005
  8. Brown, B. L.; Hadley, M.; Page, R. Protein Expr. Purif. 2008, 62, 9. https://doi.org/10.1016/j.pep.2008.07.003
  9. De Marco, V.; Stier, G.; Blandin, S.; de Marco, A. Biochem. Biophys. Res. Commun. 2004, 322, 766. https://doi.org/10.1016/j.bbrc.2004.07.189
  10. Zhang, Y. B.; Howitt, J.; McCorkle, S.; Lawrence, P.; Springer, K.; Freimuth, P. Protein Expr. Purif. 2004, 36, 207. https://doi.org/10.1016/j.pep.2004.04.020
  11. Malakhov, M. P.; Mattern, M. R.; Malakhova, O. A.; Drinker, M.; Weeks, S. D.; Butt, T. R. J. Struct. Funct. Genomics 2004, 5, 75.
  12. Volonte, F.; Marinelli, F.; Gastaldo, L.; Sacchi, S.; Pilone, M. S.; Pollegioni, L.; Molla, G. Protein Expr. Purif. 2008, 61, 131. https://doi.org/10.1016/j.pep.2008.05.010
  13. Bessette, P. H.; Aslund, F.; Beckwith, J.; Georgiou, G. Proc. Natl. Acad Sci. USA 1999, 96, 13703. https://doi.org/10.1073/pnas.96.24.13703
  14. Brown, B. L.; Grigoriu, S.; Kim, Y.; Arruda, J. M.; Davenport, A.; Wood, T. K.; Peti, W.; Page, R. PLoS Pathogens 2009, 5, e1000706. https://doi.org/10.1371/journal.ppat.1000706
  15. Wang, Y. H.; Ayrapetov, M. K.; Lin, X.; Sun, G. Biochem. Biophys. Res. Commun. 2006, 346, 606. https://doi.org/10.1016/j.bbrc.2006.05.180
  16. Chen, Y.; Song, J.; Sui, S. F.; Wang, D. N. Protein Expr. Purif. 2003, 32, 221. https://doi.org/10.1016/S1046-5928(03)00233-X
  17. Sahu, S. K.; Rajasekharan, A.; Gummadi, S. N. Biotechnol. Lett. 2009, 31, 1745. https://doi.org/10.1007/s10529-009-0073-7
  18. Hoffmann, F.; Rinas, U. In Physiological Stress Responses in Bioprocesses; Springer Berlin/Heidelberg: 2004; Vol. 89, p 73.
  19. Chae, Y. K.; Cho, K. S.; Chun, W.; Lee, K. Protein Pept. Lett. 2003, 10, 369. https://doi.org/10.2174/0929866033478861
  20. Chae, Y. K.; Moon, W. J.; Cho, J. Y. Protein Expr. Purif. 2009, 65, 267. https://doi.org/10.1016/j.pep.2009.02.001
  21. Chae, Y. K.; Lee, W. Bull. Korean Chem. Soc. 2008, 29, 2449. https://doi.org/10.5012/bkcs.2008.29.12.2449
  22. Chan, E. C.; Koh, P. K.; Mal, M.; Cheah, P. Y.; Eu, K. W.; Backshall, A.; Cavill, R.; Nicholson, J. K.; Keun, H. C. J. Proteome Res. 2009, 8, 352. https://doi.org/10.1021/pr8006232
  23. Lewis, I. A.; Schommer, S. C.; Hodis, B.; Robb, K. A.; Tonelli, M.; Westler, W. M.; Sussman, M. R.; Markley, J. L. Anal. Chem. 2007, 79, 9385. https://doi.org/10.1021/ac071583z
  24. Lewis, I. A.; Schommer, S. C.; Markley, J. L. Magn. Reson. Chem. 2009, 47(Suppl 1), S123. https://doi.org/10.1002/mrc.2526
  25. Fox, B. G.; Blommel, P. G. In Current Protocols in Protein Science; John Wiley & Sons, Inc.: 2001, p Unit 5.13.
  26. Robinette, S. L.; Ajredini, R.; Rasheed, H.; Zeinomar, A.; Schroeder, F. C.; Dossey, A. T.; Edison, A. S. Anal. Chem. 2011, 83, 1649. https://doi.org/10.1021/ac102724x
  27. Motta, A.; Paris, D.; Melck, D. Anal. Chem. 2010, 82, 2405. https://doi.org/10.1021/ac9026934
  28. Martineau, E.; Giraudeau, P.; Tea, I.; Akoka, S. J. Pharm. Biomed. Anal. 2011, 54, 252. https://doi.org/10.1016/j.jpba.2010.07.046
  29. Lewis, I. A.; Schommer, S. C.; Hodis, B.; Robb, K. A.; Tonelli, M.; Westler, W. M.; Suissman, M. R.; Markley, J. L. Anal. Chem. 2007, 79, 9385. https://doi.org/10.1021/ac071583z
  30. Tkachenko, A.; Nesterova, L.; Pshenichnov, M. Archives of Microbiology 2001, 176, 155. https://doi.org/10.1007/s002030100301

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