Magnetic Immobilization of Dispersin B with Activity in Degradation of Bacterial Biofilm

Dispersin B (DspB) is a member of glycoside hydrolase family 20 (GH20) and catalyzes degradation of biofilms forming by pathogenic bacteria such as Staphylococcus aureus. Magnetoreceptor (MagR) is a magnetic protein that can be used as a fusion partner for functionally immobilizing proteins on magnetic surfaces. In the present study, a recombinant protein DspB-MagR was constructed by fusing MagR to the C-terminus of DspB and expressed in Escherichia coli. Magnetic immobilization of purified DspB-MagR on Fe3O4@SiO2 nanoparticles was achieved and characterized by means of various techniques including SDS-PAGE, FTIR spectrometry, TGA measurements, zeta potential measurement, and SEM analysis. Stability and activity of immobilized DspB-MagR on Fe3O4@SiO2 nanoparticles were analyzed under different conditions such as temperature, pH, and storage time. Removal of biofilms forming by Staphylococcus aureus and other medical source bacterial species were achieved by using Fe3O4@SiO2 nanoparticles loading with DspB-MagR. This work promoted potential applications of DspB and similar enzymes for medical purposes.

Magnetic Immobilization of Pichia pastoris Cells for the Production of Recombinant Human Serum Albumin

Magnetic immobilization as a novel technique was used to immobilize recombinant Pichia pastoris (GS115 Albumin) cells to produce human serum albumin (HSA). In this regard, magnetic nanoparticles (MNPs) coated with amino propyl triethoxy silane (APTES) were synthesized. P. pastoris cells were decorated with MNPs via nonspecific interactions. Decorated cells were magneto-responsible and easily harvested by applying an external magnetic field. The efficiency of magnetic immobilization (Ei) for cell removal was in direct relation with the MNP concentration and time of exposure to the magnetic field. By increasing the nanoparticles concentration, cells were harvested in a shorter period. Complete cell removal (Ei ≈ 100) was achieved in ≥0.5 mg/mL of MNPs in just 30 s. HSA is produced in an extremely high cell density (OD ~20) and it is the first time that magnetic immobilization was successfully employed for harvesting such a thick cell suspension. After 5 days of induction the cells, which were immobilized with 0.25 to 1 mg/mL of nanoparticles, showed an increased potency for recombinant HSA production. The largest increase in HSA production (38.1%) was achieved in the cells that were immobilized with 0.5 mg/mL of nanoparticles. These results can be considered as a novel approach for further developments in the P. pastoris-based system.