Cyclodextrin glucanotransferase (CGTase) from Amphibacillus
sp. NPST-10 was covalently
immobilized onto amino-functionalized magnetic double mesoporous core–shell silica nanospheres
), and the properties of the immobilized enzyme were investigated. The
synthesis process of the nanospheres included preparing core magnetic magnetite (Fe3
nanoparticles, coating the Fe3
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.
The results indicated that the enzyme immobilization by covalent attachment onto the activated
, 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.
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.