The Design

The project will use synthetic biology to create a novel fusion protein that can specifically bio-tag polyethylene terephthalate (PET) plastic, so that it can be sorted and recycled correctly. Synthetic biology is efficient, cost effective, and specific. The proteins, which are produced via a bacterial chassis called Bacillus subtilis, are created efficiently and at low cost. These proteins also provide high specificity due to a specific 3-dimensional shape that adheres selectively to PET polymers. The 4 constructs that we have designed, with the help of our mentors and previous iGem teams, include:

• a polyethylene terephthalate hydrolase (PET-ase) fused to a red fluorescent protein, (or RFP) called mCherry, which give the protein its colour aspect.
• a hydrophobin called BslA,
• a PET-ase without the RFP, and
• a BslA without RFP.

A LipA secretion tag is added to each construct to signal the bacteria to secrete the proteins out of the cell for easier purification. We chose to use this Bacillus over E. coli because of its natural ability to produce hydrophobins, and because it is better at secreting proteins than other bacteria. Bacillus is also naturally occurring in the environment, and has reduced risk for environmental contamination concerns.

The PET-ase is an enzyme that naturally binds to PET plastic, and the mCherry RFP it is paired with will visually indicate when the protein has adhered. The hydrophobin is “water-fearing” and will therefore bind to several surfaces. However, for this project, it will be used to help adhere the PET-ase specifically to PET plastic. We are using the four proteins in combination with each other and test their effectiveness at tagging PET plastic.

Implemented prototype, what could be seen in a sorting facility.

Machine Prototype

With our constructs design in place, we had to design a way of using them in a real life situation. Drawing on our experiences visiting real sorting facilities, and using the feedback and insights gained from the people working in this industry, we have designed a prototype using existing technology to adapt to our solution. A simplified description of our prototype includes the following steps:

1. Incoming, unsorted plastics move along a conveyor belt and pass through a bath of our purified protein bio-tag.
2. Our bio-tag selectively adheres only to PET plastics.
3. Next all plastics will pass through a wash or rinse. The bio-tag is removed from any non-PET plastics.
4. An optical scanner detects the fluorescent signature of mCherry on the PET plastics, and will separate it from the rest of the plastic.
5. In future, similar bio-tags can be developed to selectively mark all other recyclable plastics using similar design principles.