Mechanoenzymatic Oxidation of LDPE via Designer Secretion System

• Arnav Bedekar, Chemical Engineering, University of Delaware

• Mark Blenner, Chemical and Biomolecular Engineering, University of Delaware
• Kevin Solomon, Chemical and Biomolecular Engineering, University of Delaware

Plastics are an ever growing problem in the modern world because they pose a threat to both the environment and human health. Many current plastic breakdown methods are impractical for long-term plastic management. Biocatalytic methods avoid many of the problems that traditional methods face, incorporating inherently green, benign chemistry with low energy requirements. We are investigating enzymatic oxidation to modify and break down Low Density Polyethylene (LDPE) into components that can be readily processed by organisms for upcycling. Previous work in our lab demonstrated that Dye Decolorizing Peroxidases (DyPs) are capable of LDPE oxidation. However, said oxidation is weak and DyP engineering efforts have to date only demonstrated limited gains in activity. We hypothesize modified conditions can improve activity. Mechanoenzymatic ball milling represents a promising route toward improved conditions for plastic oxidation. This technique mimics the mechanical stimulation and high concentration of these enzymes in their native environments that may allow for more plastic oxidation with our engineered DyPs. However, these conditions require large quantities of enzyme. Rather than needing to grow lyse, and purify large quantities of protein from Escherichia coli, we can leverage the modified type III secretion system using the host, Salmonella enterica. We demonstrate secretion of a prototypical DyP and successful lyophilization of a DyP without loss of activity using the Reactive Black 5 decolorization assay. By combining the oxidative ability of DyPs with the grinding activity of ball milling, a synergistic effect is expected to be achieved which will lead to enhanced oxidation. We aim to measure oxidation and degradation of mechanoenzymatically milled LDPE by Fourier Transform Infrared Spectroscopy and High Temperature Gel Permeation Chromatography respectively. Future work will focus on optimizing both the DyP and milling conditions to maximize oxidation.