Covalent immobilization of peanut β-amylase for producing industrial nano-biocatalysts: A comparative study of kinetics, stability and reusability of the immobilized enzyme

Stability of enzymes is an important parameter for their industrial applicability. Here, we report successful immobilization of β-amylase (bamyl) from peanut (Arachis hypogaea) onto Graphene oxide-carbon nanotube composite (GO-CNT), Graphene oxide nanosheets (GO) and Iron oxide nanoparticles (Fe3O4). The Box-Behnken Design of Response Surface Methodology (RSM) was used which optimized parameters affecting immobilization and gave 90%, 88% and 71% immobilization efficiency, respectively, for the above matrices. β-Amylase immobilization onto GO-CNT (bamyl@GO-CNT) and Fe3O4 (bamyl@Fe3O4), resulted into approximately 70% retention of activity at 65 °C after 100 min of exposure. We used atomic force microscopy (AFM), scanning and transmission electron microscopy (SEM and TEM), Fourier transformed infrared (FT-IR) spectroscopy and fluorescence microscopy for characterization of free and enzyme bound nanostructures (NS). Due to the non-toxic nature of immobilization matrices and simple but elegant immobilization procedure, these may have potential utility as industrial biocatalysts for production of maltose.