α-Chymotrypsin Immobilized on a Low-Density Polyethylene Surface Successfully Weakens Escherichia coli Biofilm Formation
- Catto, Cristina [ Department of Food, Environmental and Nutritional Sciences, Università degli Studi di Milano ]
- Secundo, Francesco [ Institute of Chemistry of Molecular Recognition, National Research Council ]
- James, Garth [ Montana State University: Center for Biofilm Engineering ]
- Villa, Federica [ Department of Food, Environmental and Nutritional Sciences, Università degli Studi di Milano ]
- Cappitelli, Francesca [ Department of Food, Environmental and Nutritional Sciences, Università degli Studi di Milano ]
The protease α-chymotrypsin (α-CT) was covalently immobilized on a low-density polyethylene (LDPE) surface, providing a new non-leaching material (LDPE-α-CT) able to preserve surfaces from biofilm growth over a long working timescale. The immobilized enzyme showed a transesterification activity of 1.24 nmol/h, confirming that the immobilization protocol did not negatively affect α-CT activity. Plate count viability assays, as well as confocal laser scanner microscopy (CLSM) analysis, showed that LDPE-α-CT significantly impacts Escherichia coli biofilm formation by (i) reducing the number of adhered cells (−70.7 ± 5.0%); (ii) significantly affecting biofilm thickness (−81.8 ± 16.7%), roughness (−13.8 ± 2.8%), substratum coverage (−63.1 ± 1.8%), and surface to bio-volume ratio (+7.1 ± 0.2-fold); and (iii) decreasing the matrix polysaccharide bio-volume (80.2 ± 23.2%). Additionally, CLSM images showed a destabilized biofilm with many cells dispersing from it. Notably, biofilm stained for live and dead cells confirmed that the reduction in the biomass was achieved by a mechanism that did not affect bacterial viability, reducing the chances for the evolution of resistant strains.