Enhanced Stability of β-Agarase Immobilized on Streptavidin-Coated Fe3O4 Nanoparticles: Effect of Biotin Linker Length

This study presents a simple method to immobilize beta-agarase onto amino-modified magnetic nanoparticles (MNPs) by biotin/streptavidin (BT/SA) recognition. The effect of BT functionalized with different lengths of poly(ethylene glycol) (PEG) on the activity and stability of heterogeneous biocatalysts was also investigated. The results showed that the enzymatic activity was retained above 93% under the immobilization conditions (pH 7.0, 25 degrees C, 6 h, and beta-agarase concentration of 1 mg/mL). The optimal pH and temperature for both free and immobilized beta-agarases were 7 and 30 degrees C, respectively. PEG length played an important role in the catalytic performance of the immobilized enzyme. As the PEG length increased, an increased trend in activity retention and yield of neoagarobiose was observed. Ultimately, the introduction of PEG(24) improved the activity retention of immobilized beta-agarase up to 135.67%, and retained 84.18 and 37.04% of residual activity after incubation for 6 h at 40 and 45 degrees C, respectively, while only 30.50 and 0% of residual activities were observed for free beta-agarase at these temperatures, indicating the significantly enhanced stability of the beta-agarase after immobilization. Additionally, beta-agarase-BTPEG24-SA@MNPs showed the highest yield of neoagarobiose at 45 degrees C, which was 1.89-fold higher than that of the free enzyme. These results indicated that immobilization of beta-agarase on BTPEG-SA@MNPs improved both the catalytic efficiency and stability of the enzyme, thereby promoting the continuous production of neoagarobiose.