Our team is located in San Diego, California, where the beach and outdoors are a major part of our lives. Many members of our team spend their free time outdoors, and choose to enjoy the nature around them. However, regardless of the location, we all notice something that intrudes on the beauty around us: plastic waste. The beach is often littered with plastic bottles or bags, even in areas with a lot of people. On the roads we take to school we see trash around us. Even in our own neighborhoods, we see plastic waste everywhere. While looking further into this issue, we found a specific plastic, polyethylene terephthalate (PET), which has contributed to a majority of plastic pollution. The National Renewable Energy Laboratory (NREL) finds that 82 million tons of PET is manufactured every year, and PET plastics are one of the largest sources of plastic waste. The NREL finds that most methods used for recycling PET plastics are inefficient and result in less profitable products [1]. We wanted to play an active part in developing solutions to degrade the plastics, and reduce plastic waste in our community. While researching, we found that PET is degraded through hydrolysis, and during this process terephthalic acid (TPA) is generated. Furthermore, we found that several microorganisms hold the capabilities to degrade PET, because of the specific enzymes found inside these microorganisms [2]. However, we realized there isn’t an efficient way to measure the amount of PET degraded. We decided we wanted to focus on a biosensor that could quantify plastic degradation. Since the amount of TPA generated is proportional to the amount of plastic degraded, we decided to develop a plasmid that can detect TPA. In the presence of TPA, this plasmid will create a fluorescence signal that can be detected. Developing this biosensor will allow scientists to measure how much PET is being degraded, which will allow them to measure and improve efficiency. We hope our project works towards educating others about the effects of PET plastic and is a step forward in solving the problem of plastic pollution.
Our project involves turning to biosensors as a complementary tool to conventional enzyme-directed evolution approaches in scientists’ quest to improve the detection and quantification of terephthalic acid (TPA) produced during the enzymatic breakdown of polyethylene terephthalate (PET). The complex molecular mechanism of cells, where genes alone lie inactive unless coordinated by other biological components, is used by biosensors. An importer protein (mucK) for TPA transport, a transcription factor (tpaR) sensitive to TPA binding, and the mScarlet-I and GFP reporter genes under control of tpaR-responsive promoters for signal detection make up a TPA biosensor system that has been designed in this context [3]. TPA binding to tpaR causes conformational changes that activate the transcription of mScarlet as a result of the process. The goal is to develop a more reliable biosensor with a wider dynamic range. Additionally, to increase sensitivity and specificity of the biosensor system, we are exploring changes to: the tpaR-responsive promoter (tpaAp), as the reporter genes, GFP and mScarlet, and trying multiple TPA importers. Previous reports have identified mutations in mucK that increase its ability to import TPA; based on this, we may explore mutagenesis studies to try to further increase mucK import efficiency and use our optimized biosensor to screen mutant libraries.These efforts are greatly aided by computational modeling and molecular docking simulations, which aid in predicting interactions and guide subsequent mutagenesis experiments. Researchers want to create a highly effective TPA biosensor that could change the monitoring of PET plastic deterioration in cellular systems by methodically examining these elements and their interactions.
The main goal of our project is to design a biosensor to monitor PET hydrolysis through terephthalic acid (TPA) detection that can be used by researchers to accurately quantify plastic degradation. We noticed that there were not accurate and reliable sensors for scientists to use. Thus, we decided to focus our project mainly on the detection of PET degradation.
When we surveyed people, we realized that not many people knew about the methods of plastic degradation or its impact on the environment. Through our social media platforms, we have made it our goal to increase awareness of plastic in the general community.
We believe it is extremely important to educate, inspire, and enable young students in the field of biology. We conducted two sessions of 5-day intro to synthetic biology and forensics summer camp for 1th-5th graders to hopefully spark their interest in the area.
Improving science communication has also been a significant tenet, especially during our iGEM season. We held a symposium to accomplish this goal of ours and invited experts of PET and plastic degradation to provide valuable feedback on our project and learn about potential strategies we could implement to improve our research.
[1] Oct. 13, 2021 | Contact media relations Share. (n.d.). Researchers engineer microorganisms to tackle pet plastic pollution. NREL.gov. https://www.nrel.gov/news/program/2021/researchers-engineer-microorganisms-to-tackle-pet-plastic-pollution.html#:~:text=More%20than%2082%20million%20metric
[2] Khairul Anuar, N. F. S., Huyop, F., Ur-Rehman, G., Abdullah, F., Normi, Y. M., Sabullah, M. K., & Abdul Wahab, R. (2022, October 20). An overview into polyethylene terephthalate (PET) hydrolases and efforts in tailoring enzymes for improved plastic degradation. MDPI. https://doi.org/10.3390/ijms232012644
[3] Pardo, I., Jha, R. K., Bermel, R. E., Bratti, F., Gaddis, M., McIntyre, E., … Johnson, C. W. (2020). Gene amplification, laboratory evolution, and biosensor screening reveal MucK as a terephthalic acid transporter in Acinetobacter baylyi ADP1. Metabolic Engineering, 62, 260–274. https://doi.org/10.1016/j.ymben.2020.09.009
[4] A focused review on recycling and hydrolysis techniques of polyethylene ... (n.d.). https://4spepublications.onlinelibrary.wiley.com/doi/10.1002/pen.26406
[5] Nature Publishing Group. (n.d.). Nature news. https://www.nature.com/scitable/blog/bio2.0/synthetic_biology_at_home/
[6] Urbanek, A. K., Kosiorowska, K. E., & Mirończuk, A. M. (2021, November 30). Current knowledge on polyethylene terephthalate degradation by genetically modified microorganisms. Frontiers in bioengineering and biotechnology. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8669999/