{% block lead %}We ask every team to think deeply and creatively about whether their project is responsible and good for the world. Consider how the world affects your work and how your work affects the world.{% endblock %}
{% block title %}Overview{% endblock %}
{% block lead %}A comprehensive view of our impact in human practices.{% endblock %}
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<h2>Symposium and Space Village</h2>
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<h3>Symposium and Space Village</h2>
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On August 3rd, we met with three other iGEM teams from local San Diego high schools at a public library to present and discuss our projects. The teams were from Canyon Crest Academy, Del Norte High School, Westview High School, and Rancho Bernardo High School. We talked about each component of our projects, from wiki design to education to lab work, as well as the fundamental concepts of our project and the issues that we plan to address. After each team presented, members from other teams were invited to ask questions or give comments on the projects. A range of issues were addressed during the symposium, including problems in agricultural, environmental, and health-related topics.
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<h2>Contacts</h2>
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<h3>Tejas Borkar</h3>
<p><i>Our iGEM mentor who helped us conceptualize the project</i></p>
<p>
Tejas Borkar, a Ph.D. student in Germany, guided us in the initial conceptualization of our project. He suggested adding components to our fusion protein and ways we would carry out a proof of concept in the lab. He helped us brainstorm ways to enhance the human practices component of our project and what to model. We learned more about iGEM through his experience, and we learned new scientific concepts we were previously unfamiliar with, such as GST pull-down assays and nuclear localization signals.
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<h3>Dr. Tom Huxford </h3>
<p><i>A biochemistry professor at San Diego State University</i></p>
<p>
Dr. Huxford met with us on Zoom on July 22nd after we emailed him to request lab space. He helped us organize our project ideas and plan the lab procedures we would carry out. He suggested testing the viability of the bacteria after being exposed to varying amounts of UV radiation instead of using COMET assays as we had previously planned. This simplified the process while still allowing us to gather data if we got that far in the lab. He also helped us acquire lab space by reaching out to other SDSU faculty members. We gained valuable insight on how to convert literature review and theoretical ideas into feasible wet lab procedures.
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<h3>Sierra Murrell</h3>
<p><i>A Ph.D. student working in the Love Lab</i></p>
<p>
Sierra Murrell and the other Love Lab students guided us through all of the lab procedures. She helped us plan our wet lab schedule and supervised us throughout the procedures. She and the other lab students, especially Mia and Mariam, taught us how to use various pieces of lab equipment, such as the Nanodrop and the gel visualizer, as well as the techniques needed for other protocols. We learned how to carry out a transformation and gel electrophoresis. She helped us gather materials and calculate the ratios we needed for ligation and digestion. We learned the basic lab protocols necessary for future experiments.
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<h3>Dr. Uduak George</h3>
<p><i>A mathematics professor specializing in modeling at San Diego State University</i></p>
<p>
Dr. George helped us figure out how to model components of our project mathematically. She taught us about the Michaelis-Menten equation and how to use it. She also gave us access to the papers where we could find the constants. We will use what we learned from our meetings with her in the future, especially because the Michaelis-Menten equation is so versatile for biology-related mathematical modeling.
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<h3>Dr. Pablo Machuca</h3>
<p><i>An aerospace engineering professor at San Diego State University</i></p>
<p>
Dr. Machuca met with us on Zoom on September 21st to talk about aerospace applications for the UV radiation issue. He told us about the current solutions and protection against UV radiation in space, although he specializes in designing spacecraft. We learned more context about the issue in a wider range of topics and not just biology. He helped us determine some future steps we can take in order to make our ideas more effective. The notes we took during this meeting are embedded below.
{% block title %}Proposed Implementation{% endblock %}
{% block lead %}The full idea behind our project and its impact. {% endblock %}
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<h2>Proposition</h2>
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<p>
We propose utilizing this fusion protein in sunscreen, so it can be easily accessed and applied. Current forms of protection from UV radiation in sunscreen are through UV filters, such as benzophenone [<ahref="#ref0">1</a>]. Although sunscreens are still effective in protecting against UV radiation, they only mitigate UV-induced damage and don’t typically reverse already existing damage.
</p>
<p>
For our project, we decided to focus on maximizing the ability of photolyase, an enzyme found in various bacteria and plant species, to repair common types of DNA damage, particularly cyclobutane pyrimidine dimers (CPDs) and pyrimidine-pyrimidone (6-4) photoproducts, within human skin cells. We plan to combine a cell penetrating peptide (specifically the TAT peptide from the HIV virus), a nuclear localization signal (from the SV40 Large T-antigen), and either a 6-4 photolyase protein (from Arabidopsis thaliana) or a CPD photolyase protein (from zebrafish) in one fusion protein, with linker sequences between each component to allow the protein to fold properly. The cell penetrating peptide will allow the fusion protein to enter the cell through ionic interactions with the cell membrane [<ahref="#ref1">2</a>]. The TAT peptide in particular has been found to have the ability to penetrate human skin cells [<ahref="#ref2">3</a>] Then, the NLS will guide the protein to the nucleus, which will be the main location of the DNA damage. [<ahref="#ref3">4</a>]. Finally, the photolyase protein, depending on whether it is CPD photolyase or 6-4 photolyase, will repair a certain kind of DNA damage caused by UV radiation [<ahref="#ref4">5</a>]. We will be using the common linker amino acid sequence GGGGS in between the three parts [<ahref="#ref5">6</a>].
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<p>
By combining these three components, we can create a system where the protein can be produced by bacteria and enter the cell on its own. It will then be able to repair the DNA damage caused by UV radiation itself rather than merely preventing it, as opposed to the pre existing approaches to this issue. We plan to incorporate this fusion protein into sunscreen so that, in combination with preventative measures, it will protect both astronauts and people on Earth from the growing problem of UV radiation. Although sunscreens with photolyase already exist, there is not much benefit that the photolyase adds as of yet, though their existence shows that photolyase, despite not naturally occurring in humans, does not negatively affect human skin [<ahref="#ref6">7</a>].
</p>
<p>
Other proteins in human skin, such as fibronectin, have been found to be acting as natural sunscreens [<ahref="#ref7">8</a>]. However, in the case of the fusion protein, we may need to employ another protein delivery system in order to ensure that it is able to make it inside the skin cells. We may be able to experiment with using polymer matrices, including collagen hydrogels, to carry the proteins and protect them from proteolysis. Collagen is already very biocompatible, although it does degrade quickly. Collagen I monomers can form cross-linking structures [<ahref="#ref8">9</a>], and the fusion proteins can be protected from within these cross-linking structures and diffuse into the skin [<ahref="#ref9">10</a>]. Other hydrogels have been successfully integrated into sunscreens as well, particularly hyaluronic acid and tannic acid hydrogels [<ahref="#ref10">11</a>], so this could be applied to collagen hydrogels as well.
</p>
<p>
If we wanted to implement fusion proteins in sunscreen, we would likely need to test its efficacy on bacteria, and then mice. We would need to thoroughly test it to ensure that there are no harmful side effects. We would also need to test to ensure that the collagen hydrogel can effectively protect the proteins from degradation, and that the proteins would actually enter the cell. If we created a sunscreen product, it would typically take about 6 months to a year in order for it to get approved [<ahref="#ref11">12</a>].
</p>
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<h3id="ref0">References</h3>
<pid="ref1">
[1] Amar, S. K., Srivastav, A. K., Dubey, D., Chopra, D., Singh, J., & Mujtaba, S. F. (2019). Sunscreen-induced expression and identification of photosensitive marker proteins in human keratinocytes under UV radiation. Toxicology and Industrial Health, 35(7), 457-465. <ahref="https://doi.org/10.1177/0748233719862128"target="_blank">https://doi.org/10.1177/0748233719862128</a>
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<pid="ref2">
[2] Vives, E., Richard, J., Rispal, C., & Lebleu, B. (2003). TAT peptide internalization: Seeking the mechanism of entry. Current Protein & Peptide Science, 4(2), 125-132. <ahref="https://doi.org/10.2174/1389203033487306"target="_blank">https://doi.org/10.2174/1389203033487306</a>
</p>
<pid="ref3">
[3] Johnson, J. L., Lowell, B. C., Ryabinina, O. P., Stephen Lloyd, R., & McCullough, A. K. (2011). TAT-Mediated delivery of a DNA repair enzyme to skin cells rapidly initiates repair of uv-induced DNA damage. Journal of Investigative Dermatology, 131(3), 753-761. <ahref="https://doi.org/10.1038/jid.2010.300"target="_blank">https://doi.org/10.1038/jid.2010.300</a>
</p>
<pid="ref4">
[4] Kalderon, D., Roberts, B. L., Richardson, W. D., & Smith, A. E. (1984). A short amino acid sequence able to specify nuclear location. Cell, 39(3), 499-509. <ahref="https://doi.org/10.1016/0092-8674(84)90457-4"target="_blank">https://doi.org/10.1016/0092-8674(84)90457-4</a>
</p>
<pid="ref5">
[5] Liu, Z., Wang, L., & Zhong, D. (2015). Dynamics and mechanisms of DNA repair by photolyase. Physical Chemistry Chemical Physics, 17(18), 11933-11949. <ahref="https://doi.org/10.1039/c4cp05286b"target="_blank">https://doi.org/10.1039/c4cp05286b</a>
</p>
<pid="ref6">
[6] Trinh, R., Gurbaxani, B., Morrison, S. L., & Seyfzadeh, M. (2004). Optimization of codon pair use within the (GGGGS)3 linker sequence results in enhanced protein expression. Molecular Immunology, 40(10), 717-722. <ahref="https://doi.org/10.1016/j.molimm.2003.08.006"target="_blank">https://doi.org/10.1016/j.molimm.2003.08.006</a>
</p>
<pid="ref7">
[7] Miot, H. A., Miot, L. D. B., Silva, M. G., & Marques, M. E. A. (2009). Physiopathology of ultraviolet radiation. Anais Brasileiros de Dermatologia, 84(4), 335-345. <ahref="https://www.scielo.br/j/abd/a/C3kY4LS3BdqYbdjcK5hrbVf/"target="_blank">https://www.scielo.br/j/abd/a/C3kY4LS3BdqYbdjcK5hrbVf/</a>
</p>
<pid="ref8">
[8] D’Orazio, J., Jarrett, S., Amaro-Ortiz, A., & Scott, T. (2013). UV radiation and the skin. International Journal of Molecular Sciences, 14(6), 12222-12248. <ahref="https://doi.org/10.1016/j.redox.2015.04.003"target="_blank">https://doi.org/10.1016/j.redox.2015.04.003</a>
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<pid="ref9">
[9] Du, L., Fan, M., & Mei, H. (2021). Application of nanotechnology in biofilm engineering: Methods and prospects. Microbial Biotechnology, 13(5), 100098. <ahref="https://doi.org/10.1016/j.mtbio.2021.100098"target="_blank">https://doi.org/10.1016/j.mtbio.2021.100098</a>
</p>
<pid="ref10">
[10] Deming, T. J. (2007). Polypeptide materials: New synthetic methods and applications in biomaterials. Progress in Polymer Science, 32(8-9), 858-875. <ahref="https://doi.org/10.1016/j.prsogpolymsci.2007.04.001"target="_blank">https://doi.org/10.1016/j.prsogpolymsci.2007.04.001</a>
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<pid="ref11">
[11] Xu, L., Zhang, C., Zhang, Y., Yan, X., & Wang, Y. (2021). The application of biopolymers for skin repair and protection: Recent advances in biopolymer technology. International Journal of Biological Macromolecules, 182, 243-259. <ahref="https://doi.org/10.1016/j.ijbiomac.2021.09.169"target="_blank">https://doi.org/10.1016/j.ijbiomac.2021.09.169</a>
</p>
<pid="ref12">
[12] Prime Matter Labs. (n.d.). What to expect when bringing a sunscreen product to market. Prime Matter Labs. <ahref="https://www.primematterlabs.com/resources/what-to-expect-when-bringing-a-sunscreen-product-to-market"target="_blank">https://www.primematterlabs.com/resources/what-to-expect-when-bringing-a-sunscreen-product-to-market</a>
