Immobilization of α-Amylase from Anoxybacillus sp. SK3-4 on ReliZyme and Immobead Supports

Abstract

α-Amylase from Anoxybacillus sp. SK3-4 (ASKA) is a thermostable enzyme that produces a high level of maltose from starches. A truncated ASKA (TASKA) variant with improved expression and purification efficiency was characterized in an earlier study. In this work, TASKA was purified and immobilized through covalent attachment on three epoxide (ReliZyme EP403/M, Immobead IB-150P, and Immobead IB-150A) and an amino-epoxide (ReliZyme HFA403/M) activated supports. Several parameters affecting immobilization were analyzed, including the pH, temperature, and quantity (mg) of enzyme added per gram of support. The influence of the carrier surface properties, pore sizes, and lengths of spacer arms (functional groups) on biocatalyst performances were studied. Free and immobilized TASKAs were stable at pH 6.0-9.0 and active at pH 8.0. The enzyme showed optimal activity and considerable stability at 60 • C. Immobilized TASKA retained 50% of its initial activity after 5-12 cycles of reuse. Upon degradation of starches and amylose, only immobilized TASKA on ReliZyme HFA403/M has comparable hydrolytic ability with the free enzyme. To the best of our knowledge, this is the first report of an immobilization study of an α-amylase from Anoxybacillus spp. and the first report of α-amylase immobilization using ReliZyme and Immobeads as supports.

Immobilization of the β-fructofuranosidase from Xanthophyllomyces dendrorhous by Entrapment in Polyvinyl Alcohol and Its Application to Neo-Fructooligosaccharides Production

“…The strategies for enzyme immobilization are commonly classified into three groups [18]: support binding (by adsorption or covalent linkages), entrapment and cross-linking. For reactions involving the transformation of carbohydrates, covalent binding is preferred over adsorption to avoid enzyme leakage [19], but most of the commercial activated carriers are expensive [20][21][22]. Cross-linking gives rise to biocatalysts with highly concentrated enzyme activity, significant stability and low production costs due to the absence of carrier, although the recovery of activity is commonly low [14,15].…”

Modification of Immobead 150 support for protein immobilization: Effects on the properties of immobilized Aspergillus oryzae β‐galactosidase

“…Immobead (Immobead‐Epx) is useful for adsorbing or forming covalent and multipoint covalent linkages with enzymes, due to the high surface area that results from its mesoporous nature, and the possibility of modifying the epoxy groups on its surface . Immobead 150 has been employed for the immobilization of formate dehydrogenase, lipase, α‐amylase, laccase and pullulanase . Several compounds have been used for modification of epoxy groups in commercial supports aiming to evaluate different methods of immobilization.…”

“…1,14,15 Immobead 150 has been employed for the immobilization of formate dehydrogenase, lipase, a-amylase, laccase and pullulanase. [16][17][18][19][20][21][22][23][24] Several compounds have been used for modification of epoxy groups in commercial supports aiming to evaluate different methods of immobilization. Each compound provides its own characteristics to the material, such as the crosslinking of glutaraldehyde; oxidation through acid solution action to produce carboxyl groups; activation of carboxyl groups with 1,1-(3dimethylamino-propyl)-ethyl-carbodiimide hydrochloride by the reaction of nucleophilic agents on protonated carbodiimide, under slightly acid conditions, with amino groups of proteins, and addition of amine groups using ethylenediamine.…”

Nanomaterials as novel supports for the immobilization of amylolytic enzymes and their applications: A review

“…As the substrate for this enzyme class is a very large macromolecule, it creates steric hindrances in the binding with the active site of enzymes. There are also some more demerits of traditional immobilization of α-amylases such as poor yield of immobilization in the case of surface binding and the problem of accessibility of substrate for entrapped and microencapsulated enzyme [19][20][21]. In order to overcome these problems, nanoparticles (NPs) based supports can provide a solution to such problems.…”