Background: Cyclodextrin glucanotransferase (CGTase) from
Amphibacillus
sp. NPST-10 was covalently
immobilized onto amino-functionalized magnetic double mesoporous core–shell silica nanospheres
(mag@d-SiO
2@m-SiO
2-NH
2), and the properties of the immobilized enzyme were investigated. The
synthesis process of the nanospheres included preparing core magnetic magnetite (Fe
3O
4)
nanoparticles, coating the Fe
3O
4 with a dense silica layer, followed by further coating with
functionalized or non-functionalized mesoporous silica shell. The structure of the synthesized
nanospheres was characterized using TEM, XRD, and FT-IR analyses. CGTase was immobilized onto the
functionalized and non-functionalized nanospheres by covalent attachment and physical adsorption.
Results: The results indicated that the enzyme immobilization by covalent attachment onto the activated
mag@d-SiO
2@m-SiO
2-NH
2, prepared using anionic surfactant, showed highest immobilization yield
(98.1%), loading efficiency (96.2%), and loading capacity 58 μg protein [CGTase]/mg [nanoparticles])
which were among the highest yields reported so far for CGTase. Compared with the free enzyme, the
immobilized CGTase demonstrated a shift in the optimal temperature from 50°C to 50–55°C, and showed a
significant enhancement in the enzyme thermal stability. The optimum pH values for the activity of the free and
immobilized CGTase were pH 8 and pH 8.5, respectively, and there was a significant improvement in pH stability
of the immobilized enzyme. Moreover, the immobilized CGTase exhibited good operational stability, retaining
56% of the initial activity after reutilizations of ten successive cycles.
Conclusion: The enhancement of CGTase properties upon immobilization suggested that the applied
nano-structured carriers and immobilization protocol are promising approach for industrial bioprocess
for production of cyclodextrins using immobilized CGTase.
