1887

Abstract

A synthetic promoter library (SPL) for has been developed, which generalizes the approach for obtaining synthetic promoters. The consensus sequence, derived from rRNA promoters extracted from the WCFS1 genome, was kept constant, and the non-consensus sequences were randomized. Construction of the SPL was performed in a vector (pSIP409) previously developed for high-level, inducible gene expression in and . A wide range of promoter strengths was obtained with the approach, covering 3–4 logs of expression levels in small increments of activity. The SPL was evaluated for the ability to drive -glucuronidase (GusA) and aminopeptidase N (PepN) expression. Protein production from the synthetic promoters was constitutive, and the most potent promoters gave high protein production with levels comparable to those of native rRNA promoters, and production of PepN protein corresponding to approximately 10–15 % of the total cellular protein. High correlation was obtained between the activities of promoters when tested in and , which indicates the potential of the SPL for other species. The SPL enables fine-tuning of stable gene expression for various applications in .

Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.28599-0
2006-04-01
2020-04-02
Loading full text...

Full text loading...

/deliver/fulltext/micro/152/4/1011.html?itemId=/content/journal/micro/10.1099/mic.0.28599-0&mimeType=html&fmt=ahah

References

  1. Ahrné S, Nobaek S, Jeppsson B, Adlerberth I, Wold A. E, Molin G. 1998; The normal Lactobacillus flora of healthy human rectal and oral mucosa. J Appl Microbiol85:88–94[CrossRef]
    [Google Scholar]
  2. Alander M, De Smet I, Nollet L, Verstraete W, Mattila-Sandholm T, von Wright A. 1999; The effect of probiotic strains on the microbiota of the Simulator of the Human Intestinal Microbial Ecosystem (SHIME). Int J Food Microbiol46:71–79[CrossRef]
    [Google Scholar]
  3. Aukrust T, Blom H. 1992; Transformation of Lactobacillus strains used in meat and vegetable fermentations. Food Res Int25:253–261[CrossRef]
    [Google Scholar]
  4. Axelsson L. 2004; Lactic acid bacteria: classification and physiology. In Lactic Acid Bacteria: Microbiological and Functional Aspects, 3rd edn. pp 1–66 Edited by Salminen S., von Wright A., Ouwehand A.. New York: Marcel Dekker;
    [Google Scholar]
  5. Axelsson L, Ahrné S. 2000; Lactic acid bacteria. In Applied Microbial Systematics pp 365–386 Edited by Priest F. G., Goodfellow M.. The Netherlands: Kluwer Academic Publishers;
    [Google Scholar]
  6. Axelsson L, Lindstad G, Naterstad K. 2003; Development of an inducible gene expression system for Lactobacillus sakei . Lett Appl Microbiol37:115–120[CrossRef]
    [Google Scholar]
  7. Chaves A. C, Fernandez M, Lerayer A. L, Mierau I, Kleerebezem M, Hugenholtz J. 2002; Metabolic engineering of acetaldehyde production by Streptococcus thermophilus . Appl Environ Microbiol68:5656–5662[CrossRef]
    [Google Scholar]
  8. Chen Y. S, Steele J. L. 2005; Analysis of promoter sequences from Lactobacillus helveticus CNRZ32 and their activity in other lactic acid bacteria. J Appl Microbiol98:64–72[CrossRef]
    [Google Scholar]
  9. de Vos W., Simons G. 1994; Gene cloning and expression systems in lactococci. In Genetics and Biotechnology of Lactic Acid Bacteria pp 52–105 Edited by Gasson M., de Vos W.. Oxford: Chapman and Hall;
    [Google Scholar]
  10. Exterkate F. A. 1984; Location of peptidases outside and inside the membrane of Streptococcus cremoris . Appl Environ Microbiol47:177–183
    [Google Scholar]
  11. Havenith C. E. G, Seegers J. F. M. L, Pouwels P. H. 2002; Gut-associated lactobacilli for oral immunisation. Food Res Int35:151–163[CrossRef]
    [Google Scholar]
  12. Hugenholtz J, Kleerebezem M, Starrenburg M, Delcour J, Hols P, de Vos W. 2000; Lactococcus lactis as a cell factory for high-level diacetyl production. Appl Environ Microbiol66:4112–4114[CrossRef]
    [Google Scholar]
  13. Jensen P. R, Hammer K. 1998a; Artificial promoters for metabolic optimization. Biotechnol Bioeng58:191–195[CrossRef]
    [Google Scholar]
  14. Jensen P. R, Hammer K. 1998b; The sequence of spacers between the consensus sequences modulates the strength of prokaryotic promoters. Appl Environ Microbiol64:82–87
    [Google Scholar]
  15. Jensen P. R, Westerhoff H. V, Michelsen O. 1993; The use of lac -type promoters in control analysis. Eur J Biochem211:181–191[CrossRef]
    [Google Scholar]
  16. Kahala M, Palva A. 1999; The expression signals of the Lactobacillus brevis slpA gene direct efficient heterologous protein production in lactic acid bacteria. Appl Microbiol Biotechnol51:71–78[CrossRef]
    [Google Scholar]
  17. Koebmann B. J, Andersen H. W, Solem C, Jensen P. R. 2002a; Experimental determination of control of glycolysis in Lactococcus lactis . Antonie van Leeuwenhoek82:237–248[CrossRef]
    [Google Scholar]
  18. Koebmann B. J, Solem C, Pedersen M. B, Nilsson D, Jensen P. R. 2002b; Expression of genes encoding F[sub]1[/sub]-ATPase results in uncoupling of glycolysis from biomass production in Lactococcus lactis . Appl Environ Microbiol68:4274–4282[CrossRef]
    [Google Scholar]
  19. Krüger C, Hu Y, Pan Q.10 other authors 2002; In situ delivery of passive immunity by lactobacilli producing single-chain antibodies. Nat Biotechnol20:702–706[CrossRef]
    [Google Scholar]
  20. Mathiesen G, Sørvig E, Blatny J, Naterstad K, Axelsson L, Eijsink V. G. H. 2004; High-level gene expression in Lactobacillus plantarum using a pheromone-regulated bacteriocin promoter. Lett Appl Microbiol39:137–143[CrossRef]
    [Google Scholar]
  21. McCracken A, Turner M. S, Giffard P, Hafner L. M, Timms P. 2000; Analysis of promoter sequences from Lactobacillus and Lactococcus and their activity in several Lactobacillus species. Arch Microbiol173:383–389[CrossRef]
    [Google Scholar]
  22. Mercenier A, Pouwels P, Chassy B. 1994; Genetic engineering of lactobacilli, leuconostocs and Streptococcus thermophilus. In Genetics and Biotechnology of Lactic Acid Bacteria pp 252–293 Edited by Gasson M., de Vos W.. Oxford: Chapman and Hall;
    [Google Scholar]
  23. Mercenier A, Pavan S, Pot B. 2003; Probiotics as biotherapeutic agents: present knowledge and future prospects. Curr Pharm Des9:175–191[CrossRef]
    [Google Scholar]
  24. Miller J. M. 1972; Experiments in Molecular Genetics Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  25. Møretrø T, Hagen B. F, Axelsson L. 1998; A new, completely defined medium for meat lactobacilli. J Appl Microbiol85:715–722[CrossRef]
    [Google Scholar]
  26. Møretrø T, Naterstad K, Wang E, Aasen I. M, Chaillou S, Zagorec M, Axelsson L. 2005; Sakacin P non-producing Lactobacillus sakei strains contain homologues of the sakacin P gene cluster. Res Microbiol156:949–960[CrossRef]
    [Google Scholar]
  27. Neu T, Henrich B. 2003; New thermosensitive delivery vector and its use to enable nisin-controlled gene expression in Lactobacillus gasseri . Appl Environ Microbiol69:1377–1382[CrossRef]
    [Google Scholar]
  28. Pavan S, Hols P, Delcour J, Geoffroy M. C, Grangette C, Kleerebezem M, Mercenier A. 2000; Adaptation of the nisin-controlled expression system in Lactobacillus plantarum : a tool to study in vivo biological effects. Appl Environ Microbiol66:4427–4432[CrossRef]
    [Google Scholar]
  29. Pouwels P. H, Leer R. J. 1993; Genetics of lactobacilli: plasmids and gene expression. Antonie van Leeuwenhoek64:85–107
    [Google Scholar]
  30. Pouwels P. H, Leer R. J, Boersma W. J. 1996; The potential of Lactobacillus as a carrier for oral immunization: development and preliminary characterization of vector systems for targeted delivery of antigens. J Biotechnol44:183–192[CrossRef]
    [Google Scholar]
  31. Pouwels P. H, Leer R. J, Shaw M, Tielen F. D, Smit E, Martinez B, Jore J, Conway P. L, Heijne den Bak-Glashouwer M. J. 1998; Lactic acid bacteria as antigen delivery vehicles for oral immunization purposes. Int J Food Microbiol41:155–167[CrossRef]
    [Google Scholar]
  32. Sambrook J, Fritsch E. F, Maniatis T. 1989; Molecular Cloning: a Laboratory Manual, 2nd edn.. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  33. Schillinger U, Lücke F.-K. 1989; Antibacterial activity of Lactobacillus sake isolated from meat. Appl Environ Microbiol55:1901–1906
    [Google Scholar]
  34. Schultz M, Veltkamp C, Dieleman L. A, Grenther W. B, Wyrick P. B, Tonkonogy S. L, Sartor R. B. 2002; Lactobacillus plantarum 299V in the treatment and prevention of spontaneous colitis in interleukin-10-deficient mice. Inflamm Bowel Dis8:71–80[CrossRef]
    [Google Scholar]
  35. Solem C, Jensen P. R. 2002; Modulation of gene expression made easy. Appl Environ Microbiol68:2397–2403[CrossRef]
    [Google Scholar]
  36. Solem C, Koebmann B. J, Jensen P. R. 2003; Glyceraldehyde-3-phosphate dehydrogenase has no control over glycolytic flux in Lactococcus lactis MG1363. J Bacteriol185:1564–1571[CrossRef]
    [Google Scholar]
  37. Sybesma W, Starrenburg M, Kleerebezem M, Mierau I, Hugenholtz J, de Vos W. M. 2003; Increased production of folate by metabolic engineering of Lactococcus lactis . Appl Environ Microbiol69:3069–3076[CrossRef]
    [Google Scholar]
  38. Sørvig E, Mathiesen G, Eijsink V. G, Axelsson L, Grönqvist S, Naterstad K. 2003; Construction of vectors for inducible gene expression in Lactobacillus sakei and L. plantarum . FEMS Microbiol Lett229:119–126[CrossRef]
    [Google Scholar]
  39. Sørvig E, Mathiesen G, Naterstad K, Eijsink V. G, Axelsson L. 2005; High-level, inducible gene expression in Lactobacillus sakei and Lactobacillus plantarum using versatile expression vectors. Microbiology151:2439–2449[CrossRef]
    [Google Scholar]
  40. van de Guchte M, Kok J, Venema G. 1991; Distance-dependent translational coupling and interference in Lactococcus lactis . Mol Gen Genet227:65–71[CrossRef]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.28599-0
Loading
/content/journal/micro/10.1099/mic.0.28599-0
Loading

Data & Media loading...

Most cited this month

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error