Polyethylene terephthalate (PET) is a form of polyester a light plastic most frequently used for food and beverage packaging, and in fibers for clothing. Since its invention in the 1940’s, PET has emerged as the most widely used plastic polymer in the world, with an annual production rate of nearly 50 million tons (Bornscheuer et al., 2016). 

 

Taking around 450 years to fully biodegrade, PET’s prolific utilization poses a serious threat to both marine biodiversity and human health. It is estimated that humans consume around 52,000 PET particles each year, and thus, researchers have identified the urgent need to develop novel strategies to combat the bioaccumulation of PET (Cox et al., 2019).

 

Current chemical and physical treatments for PET pollution are expensive and release secondary pollutants into the environment, rendering them counterproductive. Therefore, researchers have turned to nature to find a cheaper, environmentally friendly solution to eliminating PET pollution. Biological recycling has been found to be a promising solution.

 

Biological recycling entails the secretion of enzymes by microorganisms to effectively convert PET back into its individual monomers (terephthalic acid and ethylene glycol), which can then be repurposed into new PET products. Cutinase, esterase, and lipase are all thermophilic hydrolase enzymes that can break down PET. However, their ability to be used widespread as a practical treatment for PET plastic pollution is limited, as they can only function at temperatures of 70℃ or higher (Then et al., 2015).

 

Fortunately, in 2016, Yoshida et al., serendipitously identified Ideonella sakaiensis as a bacteria that evolved naturally to break down PET as a source of carbon by secreting IsPETase. Not only does IsPETase have a higher degradation efficiency than cutinase, esterase, lipase, and the other thermophilic hydrolase enzymes, but it also functions at moderate temperatures, making it a promising, environmentally applicable enzyme.

 

Although IsPETase exhibits the highest degradation efficiency at moderate temperatures among the enzymes tested thus far, its ability to function under the harsh stresses of the extracellular environment must be improved before its widespread application can be realized. It takes 96 hours for IsPETase to completely degrade 4 mm of square PET film, which is not nearly efficient enough for widespread use (Knott et at., 2020). In order to address this gap in literature, both the thermal stability and the degradation efficiency of IsPETase must be improved.

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In our research, we hypothesized that by utilizing the novel Protein Repair One-Stop Shop (PROSS) computational algorithm, we would be able to develop IsPETase variants with higher thermal stability than wild-type IsPETase.

 

Over two years, using PROSS, we generated variants with some of the highest melting point temperatures ever reported, and we were able to generate huge improvements with an incredibly efficient process.

 

The bioremediation of plastic pollution presents a novel approach to protecting our oceans from plastic pollution and conserving biodiversity. The insight gathered in our research about protein structures that enhance the thermal properties of IsPETase will be utilized to develop a sustainable, cost-effective solution to plastic waste reduction. The information gained about IsPETase mutations in this study will be extremely useful in future studies involving the redesign of IsPETase for optimization of its functionality. Our research will contribute toward solving plastic pollution via the complete biodegradation of polyethylene terephthalate. 

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