Team:Stanford-Brown/SB16 BioMembrane UV
The Problem
The high energy content of ionizing radiation in space poses a severe threat to life, technology, and synthetic materials alike in an extraterrestrial context. Energized subatomic particles generated from nuclear events carry over large distances in space, and are capable of tearing through organic materials and altering molecular bonds. On Earth, the vast majority of space radiation is deflected by the magnetosphere. Much of the radiation that passes through Earth’s magnetic field is then filtered out by the atmosphere, allowing only UV, visible light, and some infrared wavelengths to reach the surface. Extended UV exposure can disrupt the molecular interactions within thin membranes, creating micropores that decrease the material’s structural integrity and impermeability.
Balloons flown in Earth’s stratosphere are offered limited UV protection due to thinning of the atmosphere with increasing altitude. Chief Scientist of World View Enterprises Alan Stern stated in a meeting with our team that the materials used in his near-space exploration vessels cannot be reused due to the radiation damage they sustain during flight. Representatives of other major ballooning projects such as Google X’s Project Loon and the Stanford Space Initiative’s Balloon team expressed to members of our team that radiation exposure was the most significant factor in limiting the lifespans of their balloons. [1] The unmet need for reusability of balloon materials flown in high altitude conditions generates a lack of sustainability that our team aimed to tackle through UV resistance.
On Earth, the main source of concern for radiation damage is ultraviolet radiation (UVR). These wavelengths are between 290 and 400 nm when they hit the Earth’s surface, and do more damage the shorter they get. UV wavelengths between 320 and 400 nm are called UV-A, and are the culprits behind sunburns and cataract formation. UV-A radiation creates indirect DNA damage through the generation of reactive oxygen species (ROS), forming strand breaks in DNA and DNA-protein crosslinks. [2] Shorter wavelengths from 290 to 320 nm are called UV-B wavelengths. UV-B is absorbed directly into DNA and causes damage to DNA at the molecular level. [3] Excessive UV-B exposure can induce significant DNA mutation and cell death, and our bioballoon will be exposed to dangerous amounts of radiation due to its high elevation during flight. Ideally, our balloon coating would have included an interweaving of zinc oxide, a white pigment that excels at blocking and reflecting UV-A and UV-B rays. [4] Zinc oxide is used in many commercial sunscreens, as a solution with as low as 25% zinc oxide content results in protection of at least SPF 20. [5] However, we wanted to produce our UV protector entirely biologically rather than attempting to incorporate an inorganic compound into our membrane. As such, we decided to focus on creating a method of UV protection for our balloon that utilises a melanin coating attached to the balloon’s outer membrane.
References
1. How to Protect Astronauts from Space Radiation on Mars. (2016). NASA. Retrieved 9 October 2016, from http://www.nasa.gov/feature/goddard/real-martians-how-to-protect-astronauts-from-space-radiation-on-mars
2. Brenner, M. & Hearing, V. (2007). The Protective Role of Melanin Against UV Damage in Human Skin†. Photochemistry And Photobiology, 84(3), 539-549. http://dx.doi.org/10.1111/j.1751-1097.2007.00226.x
3. Ultraviolet Radiation: How It Affects Life on Earth : Feature Articles. (2016). Earthobservatory.nasa.gov. Retrieved 9 July 2016, from http://earthobservatory.nasa.gov/Features/UVB/
4. Soni, R. & Toetia, M. (2016). Impact of Zinc Oxide on the UV Absorbance and Mechanical Properties of UV cured films. Presentation, Department of Chemistry, C.C.S. University of Meerut.
5. ZINC OXIDE | ZnO - PubChem. (2016). Pubchem.ncbi.nlm.nih.gov. Retrieved 9 July 2016, from https://pubchem.ncbi.nlm.nih.gov/compound/zinc_oxide#section=Therapeutic-Uses
References
1. How to Protect Astronauts from Space Radiation on Mars. (2016). NASA. Retrieved 9 October 2016, from http://www.nasa.gov/feature/goddard/real-martians-how-to-protect-astronauts-from-space-radiation-on-mars
2. Brenner, M. & Hearing, V. (2007). The Protective Role of Melanin Against UV Damage in Human Skin†. Photochemistry And Photobiology, 84(3), 539-549. http://dx.doi.org/10.1111/j.1751-1097.2007.00226.x
3. Ultraviolet Radiation: How It Affects Life on Earth : Feature Articles. (2016). Earthobservatory.nasa.gov. Retrieved 9 July 2016, from http://earthobservatory.nasa.gov/Features/UVB/
4. Soni, R. & Toetia, M. (2016). Impact of Zinc Oxide on the UV Absorbance and Mechanical Properties of UV cured films. Presentation, Department of Chemistry, C.C.S. University of Meerut.
5. ZINC OXIDE | ZnO - PubChem. (2016). Pubchem.ncbi.nlm.nih.gov. Retrieved 9 July 2016, from https://pubchem.ncbi.nlm.nih.gov/compound/zinc_oxide#section=Therapeutic-Uses
Our Solution
To install radiation resistance in our biomembranes, the team investigated the absorptive properties of melanin and designed a novel binding mechanism to incorporate the pigment directly into materials. Melanin is the primary source of naturally-occurring UV protection in human skin. In response to human skin damage through UV exposure, keratinocytes release cytokines to stimulate melanocytes’ production of melanin, which functions as a UV absorbent and generates the phenotypic response of darker skin. [1] Melanin when used as a sunscreen has been shown to absorb between 50-75% of UVR. [2] Two main types of melanin exist: eumelanin and pheomelanin, with eumelanin being darker and browner in color and more effective at photoprotection. After significant UV-A absorbance, melanin tends to produce ROS and create single-strand DNA breaks, with pheomelanin being significantly more susceptible to photodegradation than eumelanin. Thus, brown eumelanin was chosen as a good candidate for our balloon’s UV protection.
References
1. Miller, A.J & Tsao, H, 2009, ‘New insights into pigmentary pathways and skin cancer’, British Journal of Dermatology, 162(1):22-28. http://www.medscape.com/viewarticle/720364
2. Brenner, M. & Hearing, V. (2007). The Protective Role of Melanin Against UV Damage in Human Skin†. Photochemistry And Photobiology, 84(3), 539-549. http://dx.doi.org/10.1111/j.1751-1097.2007.00226.x
1. Miller, A.J & Tsao, H, 2009, ‘New insights into pigmentary pathways and skin cancer’, British Journal of Dermatology, 162(1):22-28. http://www.medscape.com/viewarticle/720364
2. Brenner, M. & Hearing, V. (2007). The Protective Role of Melanin Against UV Damage in Human Skin†. Photochemistry And Photobiology, 84(3), 539-549. http://dx.doi.org/10.1111/j.1751-1097.2007.00226.x
Melanin Production
Melanin is synthesized by many biological organisms from the aromatic amino acid L-tyrosine through the activity of tyrosinases. These enzymes catalyze the hydroxylation of L-tyrosine to L-DOPA, as well as a further oxidation to produce the cyclical dopachrome. Eumelanin is formed in vivo by the non-enzymatic oxidation and polymerization of dopachrome. [1] The MutmelA gene endogenous to Rhizobium etli encodes a feedback resistant tyrosinase that has been shown to be effective when transfected into E. coli. [2] Based on the work of MI Chávez-Béjar, our team sought to increase the production of L-tyrosine in E. coli through metabolic engineering to shuttle the flow of carbon from the cells’ central metabolism into the shikimate pathway.
Increasing L-tyrosine Levels
L-tyrosine, along with many other common aromatic compounds, is produced endogenously in E. coli through the shikimate pathway. In this pathways initial reaction, DAHP synthase catalyzes the conversion of Phosphoenolpyruvic acid (PEP) and Erythrose 4-phosphate (E4P) to DAHP. PEP and E4P are naturally derived from glucose, and are present at high concentrations in the cytoplasm of E. coli grown in glucose containing medium. E. coli naturally contain three isoenzymes of DAHP synthase, each of which are inhibited by the increasing concentrations of specific amino acids produced via their activity. This negative feedback limits the production capacity of L-tyrosine in wild type E. coli. To upregulate L-tyrosine production, we investigated and biobricked multiple feedback resistant variants of the DAHP synthase isoenzymes AroG (BBa_K2027013, BBa_K2027014) and AroF (BBa_K2027008).
L-tyrosine, along with many other common aromatic compounds, is produced endogenously in E. coli through the shikimate pathway. In this pathways initial reaction, DAHP synthase catalyzes the conversion of Phosphoenolpyruvic acid (PEP) and Erythrose 4-phosphate (E4P) to DAHP. PEP and E4P are naturally derived from glucose, and are present at high concentrations in the cytoplasm of E. coli grown in glucose containing medium. E. coli naturally contain three isoenzymes of DAHP synthase, each of which are inhibited by the increasing concentrations of specific amino acids produced via their activity. This negative feedback limits the production capacity of L-tyrosine in wild type E. coli. To upregulate L-tyrosine production, we investigated and biobricked multiple feedback resistant variants of the DAHP synthase isoenzymes AroG (BBa_K2027013, BBa_K2027014) and AroF (BBa_K2027008).
References
1. García-Borrón, J. C. and Solano, F. (2002), Molecular Anatomy of Tyrosinase and its Related Proteins: Beyond the Histidine-Bound Metal Catalytic Center. Pigment Cell Research, 15: 162–173. doi:10.1034/j.1600-0749.2002.02012.x
2. María I Chávez-Béjar, Metabolic engineering of Escherichia coli to optimize melanin synthesis from glucose. Microbial Cell Factories, 2013 Nov 13;12:108. doi: 10.1186/1475-2859-12-108.
1. García-Borrón, J. C. and Solano, F. (2002), Molecular Anatomy of Tyrosinase and its Related Proteins: Beyond the Histidine-Bound Metal Catalytic Center. Pigment Cell Research, 15: 162–173. doi:10.1034/j.1600-0749.2002.02012.x
2. María I Chávez-Béjar, Metabolic engineering of Escherichia coli to optimize melanin synthesis from glucose. Microbial Cell Factories, 2013 Nov 13;12:108. doi: 10.1186/1475-2859-12-108.
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Binding Agent
Coating a balloon with a protective agent requires strict homogeneity to be useful. Kirill Safin, the team lead of Stanford Space Initiative’s Ballooning project, provided insight into the adverse effects of uneven coatings on balloons. Irregular coatings can shorten balloon lifespans compared to uncoated balloons by creating inconsistencies in a membranes tensile properties and allowing the material to stretch more in one location than another. In balloons with flexible membranes, changes in ambient pressure and temperature lead to fluctuations in volume that stretch the balloons’ membranes. This can cause an inelastic coating to become uneven, exposing weak spots susceptible to tearing.
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Results
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Conclusion
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