Team:ETH Zurich/Interplay

From 2012.igem.org

(Difference between revisions)
Line 29: Line 29:
=== PABA and the Sun protection factor ===
=== PABA and the Sun protection factor ===
-
Last but not least we asked ourselves how much PABA we need to produce to achieve a certain Sun Protection Factor (SPF). Or, vice versa, given the amount of PABA our bacteria produced, what is the achieved Sun Protection Factor. This helped to decide for a chorismate accumulating strain to produce more PABA to yield a higher SPF. It also reveiled that we also need an UV-A absorbing molecule such as the PABA related [http://en.wikipedia.org/wiki/Padimate_O Padimate O] to reach meaningful SPFs since UV-A and UV-B radiation are likewise dangerous.
+
Last but not least we asked ourselves how much [[Team:ETH_Zurich/PABA|PABA]] we need to produce to achieve a certain [[Team:ETH_Zurich/Modeling/SPF_model|Sun Protection Factor]] (SPF). Or, vice versa, given the amount of PABA our bacteria produced, what is the achieved Sun Protection Factor. This helped to decide for a chorismate accumulating strain to produce more PABA to yield a higher SPF. It also reveiled that we also need an UV-A absorbing molecule such as the PABA related [http://en.wikipedia.org/wiki/Padimate_O Padimate O] to reach meaningful SPFs since UV-A and UV-B radiation are likewise dangerous.
{{:Team:ETH_Zurich/Templates/Footer}}
{{:Team:ETH_Zurich/Templates/Footer}}

Revision as of 13:15, 23 October 2012

Eth ecolipseeth logo.png
Eth igem logo.png

Contents

Interaction between the laboratory and the modelling

Lab and Modelling, or Modelling in the Lab?

To save time and work, laboratory work and in silico modelling should interact closely so that the model gives useful insights and the laboratory provides results from experimental set ups to refine the model. Here we want to outline how laboratory work and modelling interacted throughout our project.

Photoinduction

Given the four available light receptors available as BioBricks, LovTap, Cph8, YcgE/F and Ccas, we wanted to know which one is best suitable for our purpose of detecting sun light. Thus we modeled the activating rates of the different receptors when exposed to sunlight, the photoinduction. This helped us to rule out YcgE/F and Ccas that did not give the desired activation under sunlight. We continued with LovTap and Cph8 for our decoder.

The photoinduction model was also used to derive the activity of our new fusion protein, UVR8-TeTRDBD to get a grasp on its activation due to sunlight.

Circuit modeling

UVR8 Circuit

We modeled the direct UV-B detection by UVR8-TeTRDBD circuit with the downstream components: the PABA producing enzymes and the pigment to get an idea how much PABA the system can produce at steady state.

UVR8 Feedback

We were interested in how the absorbtion of UV-B radiation by our protection molecule PABA can affect the activation of our UVR8-TeTRDBD fusion protein. The negative feedback reduces the output, but has no other adverse effects on the system.

Decoder Circuit LovTap Cph8

To see how we can indirectly detect sunlight with blue and red light receptors, we came up with the idea and biological implementation of the decoder. Our model showed that the decoder can theoretically work and resulted in constraints given the promotor strength in the system.

PABA and the Sun protection factor

Last but not least we asked ourselves how much PABA we need to produce to achieve a certain Sun Protection Factor (SPF). Or, vice versa, given the amount of PABA our bacteria produced, what is the achieved Sun Protection Factor. This helped to decide for a chorismate accumulating strain to produce more PABA to yield a higher SPF. It also reveiled that we also need an UV-A absorbing molecule such as the PABA related [http://en.wikipedia.org/wiki/Padimate_O Padimate O] to reach meaningful SPFs since UV-A and UV-B radiation are likewise dangerous.


References

  • Brown, B. a, Headland, L. R., & Jenkins, G. I. (2009). UV-B action spectrum for UVR8-mediated HY5 transcript accumulation in Arabidopsis. Photochemistry and photobiology, 85(5), 1147–55.
  • Christie, J. M., Salomon, M., Nozue, K., Wada, M., & Briggs, W. R. (1999): LOV (light, oxygen, or voltage) domains of the blue-light photoreceptor phototropin (nph1): binding sites for the chromophore flavin mononucleotide. Proceedings of the National Academy of Sciences of the United States of America, 96(15), 8779–83.
  • Christie, J. M., Arvai, A. S., Baxter, K. J., Heilmann, M., Pratt, A. J., O’Hara, A., Kelly, S. M., et al. (2012). Plant UVR8 photoreceptor senses UV-B by tryptophan-mediated disruption of cross-dimer salt bridges. Science (New York, N.Y.), 335(6075), 1492–6.
  • Cloix, C., & Jenkins, G. I. (2008). Interaction of the Arabidopsis UV-B-specific signaling component UVR8 with chromatin. Molecular plant, 1(1), 118–28.
  • Cox, R. S., Surette, M. G., & Elowitz, M. B. (2007). Programming gene expression with combinatorial promoters. Molecular systems biology, 3(145), 145. doi:10.1038/msb4100187
  • Drepper, T., Eggert, T., Circolone, F., Heck, A., Krauss, U., Guterl, J.-K., Wendorff, M., et al. (2007). Reporter proteins for in vivo fluorescence without oxygen. Nature biotechnology, 25(4), 443–5
  • Drepper, T., Krauss, U., & Berstenhorst, S. M. zu. (2011). Lights on and action! Controlling microbial gene expression by light. Applied microbiology, 23–40.
  • EuropeanCommission (2006). SCIENTIFIC COMMITTEE ON CONSUMER PRODUCTS SCCP Opinion on Biological effects of ultraviolet radiation relevant to health with particular reference to sunbeds for cosmetic purposes.
  • Elvidge, C. D., Keith, D. M., Tuttle, B. T., & Baugh, K. E. (2010). Spectral identification of lighting type and character. Sensors (Basel, Switzerland), 10(4), 3961–88.
  • GarciaOjalvo, J., Elowitz, M. B., & Strogatz, S. H. (2004). Modeling a synthetic multicellular clock: repressilators coupled by quorum sensing. Proceedings of the National Academy of Sciences of the United States of America, 101(30), 10955–60.
  • Gao Q, Garcia-Pichel F. (2011). Microbial ultraviolet sunscreens. Nat Rev Microbiol. 9(11):791-802.
  • Goosen N, Moolenaar GF. (2008) Repair of UV damage in bacteria. DNA Repair (Amst).7(3):353-79.
  • Heijde, M., & Ulm, R. (2012). UV-B photoreceptor-mediated signalling in plants. Trends in plant science, 17(4), 230–7.
  • Hirose, Y., Narikawa, R., Katayama, M., & Ikeuchi, M. (2010). Cyanobacteriochrome CcaS regulates phycoerythrin accumulation in Nostoc punctiforme, a group II chromatic adapter. Proceedings of the National Academy of Sciences of the United States of America, 107(19), 8854–9.
  • Hirose, Y., Shimada, T., Narikawa, R., Katayama, M., & Ikeuchi, M. (2008). Cyanobacteriochrome CcaS is the green light receptor that induces the expression of phycobilisome linker protein. Proceedings of the National Academy of Sciences of the United States of America, 105(28), 9528–33.
  • Kast, Asif-Ullah & Hilvert (1996) Tetrahedron Lett. 37, 2691 - 2694., Kast, Asif-Ullah, Jiang & Hilvert (1996) Proc. Natl. Acad. Sci. USA 93, 5043 - 5048
  • Kiefer, J., Ebel, N., Schlücker, E., & Leipertz, A. (2010). Characterization of Escherichia coli suspensions using UV/Vis/NIR absorption spectroscopy. Analytical Methods, 9660. doi:10.1039/b9ay00185a
  • Kinkhabwala, A., & Guet, C. C. (2008). Uncovering cis regulatory codes using synthetic promoter shuffling. PloS one, 3(4), e2030.
  • Krebs in Deutschland 2005/2006. Häufigkeiten und Trends. 7. Auflage, 2010, Robert Koch-Institut (Hrsg) und die Gesellschaft der epidemiologischen Krebsregister in Deutschland e. V. (Hrsg). Berlin.
  • Lamparter, T., Michael, N., Mittmann, F., & Esteban, B. (2002). Phytochrome from Agrobacterium tumefaciens has unusual spectral properties and reveals an N-terminal chromophore attachment site. Proceedings of the National Academy of Sciences of the United States of America, 99(18), 11628–33.
  • Levskaya, A. et al (2005). Engineering Escherichia coli to see light. Nature, 438(7067), 442.
  • Mancinelli, A. (1986). Comparison of spectral properties of phytochromes from different preparations. Plant physiology, 82(4), 956–61.
  • Nakasone, Y., Ono, T., Ishii, A., Masuda, S., & Terazima, M. (2007). Transient dimerization and conformational change of a BLUF protein: YcgF. Journal of the American Chemical Society, 129(22), 7028–35.
  • Orth, P., & Schnappinger, D. (2000). Structural basis of gene regulation by the tetracycline inducible Tet repressor-operator system. Nature structural biology, 215–219.
  • Parkin, D.M., et al., Global cancer statistics, 2002. CA: a cancer journal for clinicians, 2005. 55(2): p. 74-108.
  • Rajagopal, S., Key, J. M., Purcell, E. B., Boerema, D. J., & Moffat, K. (2004). Purification and initial characterization of a putative blue light-regulated phosphodiesterase from Escherichia coli. Photochemistry and photobiology, 80(3), 542–7.
  • Rizzini, L., Favory, J.-J., Cloix, C., Faggionato, D., O’Hara, A., Kaiserli, E., Baumeister, R., et al. (2011). Perception of UV-B by the Arabidopsis UVR8 protein. Science (New York, N.Y.), 332(6025), 103–6.
  • Roux, B., & Walsh, C. T. (1992). p-aminobenzoate synthesis in Escherichia coli: kinetic and mechanistic characterization of the amidotransferase PabA. Biochemistry, 31(30), 6904–10.
  • Strickland, D. (2008). Light-activated DNA binding in a designed allosteric protein. Proceedings of the National Academy of Sciences of the United States of America, 105(31), 10709–10714.
  • Sinha RP, Häder DP. UV-induced DNA damage and repair: a review. Photochem Photobiol Sci. (2002). 1(4):225-36
  • Sambandan DR, Ratner D. (2011). Sunscreens: an overview and update. J Am Acad Dermatol. 2011 Apr;64(4):748-58.
  • Tabor, J. J., Levskaya, A., & Voigt, C. A. (2011). Multichromatic Control of Gene Expression in Escherichia coli. Journal of Molecular Biology, 405(2), 315–324.
  • Thibodeaux, G., & Cowmeadow, R. (2009). A tetracycline repressor-based mammalian two-hybrid system to detect protein–protein interactions in vivo. Analytical biochemistry, 386(1), 129–131.
  • Tschowri, N., & Busse, S. (2009). The BLUF-EAL protein YcgF acts as a direct anti-repressor in a blue-light response of Escherichia coli. Genes & development, 522–534.
  • Tschowri, N., Lindenberg, S., & Hengge, R. (2012). Molecular function and potential evolution of the biofilm-modulating blue light-signalling pathway of Escherichia coli. Molecular microbiology.
  • Tyagi, A. (2009). Photodynamics of a flavin based blue-light regulated phosphodiesterase protein and its photoreceptor BLUF domain.
  • Vainio, H. & Bianchini, F. (2001). IARC Handbooks of Cancer Prevention: Volume 5: Sunscreens. Oxford University Press, USA
  • Quinlivan, Eoin P & Roje, Sanja & Basset, Gilles & Shachar-Hill, Yair & Gregory, Jesse F & Hanson, Andrew D. (2003). The folate precursor p-aminobenzoate is reversibly converted to its glucose ester in the plant cytosol. The Journal of biological chemistry, 278.
  • van Thor, J. J., Borucki, B., Crielaard, W., Otto, H., Lamparter, T., Hughes, J., Hellingwerf, K. J., et al. (2001). Light-induced proton release and proton uptake reactions in the cyanobacterial phytochrome Cph1. Biochemistry, 40(38), 11460–71.
  • Wegkamp A, van Oorschot W, de Vos WM, Smid EJ. (2007 )Characterization of the role of para-aminobenzoic acid biosynthesis in folate production by Lactococcus lactis. Appl Environ Microbiol. Apr;73(8):2673-81.
Retrieved from "http://2012.igem.org/Team:ETH_Zurich/Interplay"