Abstract:

Polyethylene terephthalate (PET) is one of the world’s most common thermoplastics. Its accumulation has detrimental impacts on the environment and human health. Currently, mechanical recycling is the best way to reduce PET in the environment. However, it consumes significant energy while emitting greenhouse gases, and it can only be performed a limited number of times. Biodegradation techniques are being explored to find more sustainable, efficient PET recycling methods. One such technique, enzymatic recycling, will only be feasible at an industrial scale once effective crystalline PET-degrading enzymes are developed. This project aims to design and express a PET hydrolase that exhibits the characteristics associated with stronger crystalline degradation activity — a hydrophobic, linear, and wide active site cleft, and increased thermostability. Amino acid residue mutations were introduced to the sequence of an existing hydrolase (PDB ID 7QJP) to enhance the desired characteristics. The resulting mutants were modeled using AlphaFold2 (a neural network for protein structure prediction) on Google Colab. In Schrödinger Maestro, they were then evaluated using the following features: protein structure alignment, protein preparation, docking (to a PET fragment), and ligand interaction diagrams; additionally, the cysteine mutation feature was used to identify potential disulfide bonds. Finally, DeepSTABp (deep learning software) was used to predict the melting temperatures. This resulted in 80 promising mutants with the desired characteristics. A preliminary transformation (heat shock) and expression were conducted for the top-performing mutant; it was successfully expressed, indicating that it’s a viable enzyme. Optimized induction conditions will yield enough for it (and similar mutants) to be harvested and used in an assay to confirm its degradation efficiency. This has the potential to allow large-scale implementation of enzymatic recycling, offering a more sustainable way to recycle PET plastic.

Speaker:

Mena Boggs, NCSSM/NC State University

Mena Boggs is an incoming freshman at North Carolina State University, where she plans to major in Chemical Engineering, Computer Science, or Bioinformatics. She is passionate about environmental protection, which inspired her research on plastic-degrading enzymes.