Abstract
Cytoplasm of Saccharomyces cerevisiae yeast cells contains a significant amount of desired intracellular products for both industrial utility and academic research. To recover intracellular compounds, it is necessary to break the yeast cells with high efficiency, which, under certain circumstances, requires the use of the lytic enzyme zymolyase to completely digest the cell walls. A promising strategy for zymolyase immobilization on silk fibroin (SF) was developed. SF/Fe3O4 magnetic microspheres (MMs) were constructed by solvent (ethanol)-induced self-assembly of SF surrounding Fe3O4 magnetic nanoparticles (MNs), which were synthesized by a coprecipitation method. Zymolyase was covalently bonded on the surface of the SF/Fe3O4 MMs by a photochemical cross-linking method to produce robust biocatalysts of zymolyase/SF/Fe3O4. The chemical, magnetic, and morphological properties of the MM supports and the immobilized zymolyase were investigated. Enzymolysis results demonstrated that the immobilized zymolyase showed good activity and stability for digesting yeast cell walls, and the biocatalyst can be readily recycled through convenient magnetic separation for reuse. At the optimum pH = 7.5, the immobilized zymolyase maintained 84% of the activity of the free zymolyase and retained 41% of its initial activity after four times of reuse. At unfavorable acidic pH = 4, the immobilized zymolyase retained 81% of its initial activity, while the free zymolyase showed no significant activity. Consequently, the SF/Fe3O4 MMs exhibit superior performance in terms of immobilizing enzymes, which have a good prospect in the biological application.