Team:Baltimore Bio-Crew/Description


Bio-Engineering E.Coli To Degrade Plastic and Save The Baltimore Inner Harbor

Project Description

From a tourists’ perspective, Baltimore City’s Inner Harbor seems to be a lively and enjoyable place to visit. The Inner Harbor offers various restaurants and shopping venues along the water. It has a lot of attractions, until someone looks closely at the water. The water that makes up the Inner Harbor is filled with plastic and other trash, especially after a rain. One of the recent approaches to get rid of trash in the Inner Harbor recently picked up around 1 million pounds of trash.
The main reason the Baltimore City Inner Harbor is polluted is the notorious issue of plastic pollution. Walking through Baltimore City’s streets, one can see pollution on every sidewalk and near every gutter. The plastic pollution is then transferred from the city’s streets into the city’s inner harbor via the sewers. According to data acquired by the Ocean Crusader’s Foundation, around 5.25 trillion pieces of plastic debris are currently in the ocean.
As a result of this pollution, more than 100,000 marine creatures and 1 million seabirds are found dead each year, usually due to strangulation by plastic or suffocation from ingesting the plastic. Not only are the animals in the oceans affected, but humans are affected as well. We eat tons of contaminated sea creatures each year. Some of these sea creatures contaminated with plastic have toxins such as diethylhexyl phthalate (DEHP), bromine compounds, and estrogenic chemicals. These toxins can affect the human body’s hormones, nervous system, and can cause cancer.
There have been plenty of approaches to eliminate plastic in the waterways. Mr. Trash Wheel in the Harbor removes plastic in the Inner Harbor by using rotating forks to pick up trash and place it on a conveyer belt. Another approach, Boomy McBoomface, tries to collect plastic before it ventures too far into the North Sea. However, both approaches only collect trash that is on the surface of the water and also costs thousands of dollars. This means that these approaches are not collecting microplastics and that it is also a burden when it comes to the cost.
Our goal is to use an approach that is efficient and cost-effective. After researching different biological approaches to eliminate the most common plastic in the oceans, polyethylene terephthalate (PET), we came across a Nature article that mentioned a bacterium called Ideonella sakaiensis that can degrade plastic into two benign monomers using two enzymes: chlorogenate esterase and lipase. The paper by Yoshida et. al. described how this bacterium uses PETase (chlorogenate esterase) to turn PET into MHET and using MHETase (lipase) to turn MHET into ethylene glycol and terephthalic acid. Due to the recent discovery of this bacterium and unknown precautions in a laboratory setting, our team decided to use the K-12 strain of Escherichia coli and genetically modify it to express the enzyme lipase and chlorogenate esterase. The K-12 strain of E.coli is commonly used in laboratories and is safe to work with.
Some applications for this genetically modified E.coli is by degrading plastic in laboratories that cannot be recycled, being used in a laundry filter to collect microplastic fibers or breaking down plastic in contained marine environments and turning them into benign monomers. By the end of last year’s competition, we had managed to insert the lipase gene into E.coli, but not the chlorogenate esterase gene. We confirmed that we had correctly inserted the lipase gene using colony PCR and gene sequencing, but we did not have the time to conduct additional assays, such as protein gels, to determine if the enzyme was being secreted from the bacteria. This year, we plan to redesign the chlorogenate esterase and lipase genes so that they contain the proper tags that will allow them to be detected, along with a secretion sequence to help the enzymes exit the cell. After we insert both genes into E.coli cells, we will test their functionality using protein gels and enzyme assays to make sure they can produce the plastic degrading enzymes and degrade PET plastic.