Coding

Part:BBa_K554004:Experience

Designed by: UNICAMP EMSE Brazil team   Group: iGEM11_UNICAMP-EMSE_Brazil   (2011-09-21)

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Applications of BBa_K554004

Response System Improvements

UNICAMP-EMSE Team aimed the development of a bacteria to be used as a probiotic and capable of sensing and interfering with stress-caused immune imbalances and related diseases. Instead of using retinoic acid and its pathway genes to act as an immune-suppressor of Th1 overactivation (as used by Stanford 2009 Team) we have chosen to use Interleukin 10 (IL-10). This choice was made upon scientific background describing this molecule as able to act in two important directions we considered interesting to create a immune balance: IL-10 is efficient to establish an Immunosuppression of Th1 lineage (by directly inhibiting IL-2 production by these cells (de Vries, 1995)), inhibit the production of inflammatory signals (Sabat et al, 2010) and also to redirect the immune cells differentiation toward Th2 (Elenkov et al, 1999). The special physiological relevance of this cytokine lies in the prevention and limitation of over-whelming specific and unspecific immune reactions and, in consequence, of tissue damage (Sabat et al, 2010). Additionally, it was observed that IL-10 was efficient in the treatment of chronic gut diseases associated with autoimmunity (Steidler et al., 2000) and also a bacteria, Lactococcus lactis, efficiently secreted biologically active murine IL-10 (Schotte et al., 2000). These data, linked to new delivery technologies (Huyghebaert et al 2005) indicate that IL-10 is suitable to be used as a probiotic.

The mRNA sequence of human IL10 had its expression in E. coli optimized by codon usage and suited to standard 23 through remotion of internal restriction sites. Additionally, RBS sequence was fusioned with the coding sequence. All these modifications were performed prior to sequence synthesis, which enabled us to create the part BBa_K554004.

References

Steidler L, Hans W, Schotte L, Neirynck S, Obermeier F, Falk W, Fiers W, Remaut E. Treatment of murine colitis by Lactococcus lactis secreting interleukin-10. Science. 2000 Aug 25;289(5483):1352-5.

Schotte L, Steidler L, Vandekerckhove J, Remaut E. Secretion of biologically active murine interleukin-10 by Lactococcus lactis.Enzyme Microb Technol. 2000 Dec;27(10):761-765.

Huyghebaert N, Vermeire A, Neirynck S, Steidler L, Remaut E, Remon JP. Secretion of biologically active murine interleukin-10 by Lactococcus lactis. Enzyme and Microbial Technology 27 (2000) 761–765

de Vries JE. Immunosuppressive and anti-inflammatory properties of interleukin 10.Ann Med. 1995 Oct;27(5):537-41.

Sabat R, Grütz G, Warszawska K, Kirsch S, Witte E, Wolk K, Geginat J. Biology of interleukin-10. Cytokine Growth Factor Rev. 2010 Oct;21(5):331-44. Epub 2010 Nov 5.



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