Determination of some adsorption and kinetic parameters of α-amylase onto Cu⁺²-PHEMA beads embedded column

Abstract

In order to investigate the biocatalytic properties of a-amylase on a composite cryogel matrix with immobilized metal affinity chromatography, Cu þ2 -attached poly(2-hydroxyethyl methacrylate) (Cu þ2 -PHEMA) beads, (2 mm size) were synthesized, then composite cryogel column was prepared by composing beads and PHEMA cryogels. After the preparation of Cu þ2 -PHEMA beads embedded cryogel column (Cu þ2 -BEC), some experiments were tested. Accordingly, the highest adsorption capacity (676.8 mg/g particles) of cryogels was achieved at acetate buffer of pH 5.0 with initial a-amylase concentration of 4 mg/mL. Immobilized enzyme has more stable pH range, between 6 and 7.5 than, the free one. Immobilization also increased the optimal activity from 25 to temperature range of 25-35 C. V max and K m of a-amylase were detected as 1.149 U/mg protein, and 11.6 Â 10 À1 mM, respectively. a-Amylase was utilized 35 times repeatedly without losing the productivity.

α-Amylase Immobilized Composite Cryogels: Some Studies on Kinetic and Adsorption Factors

“…The maximum adsorption capacity was occured when the pH reached at 4.0. This situation can be explained as follows: α-amylase contains acidic amino acids (i.e., aspartate and glutamate) mostly since its isoelectric point is near the acidic region [14,31]. It was noticed that IMAC matrix can act as a pseudo-anion exchanger even if there are no IMAC active amino acid residues (i.e., histidine, cysteine) near by the surface for metal ligand [32].…”

“…Immobilization of α-amylase on carriers appears the most appropriate method to acquire stable and repeated used forms of enzymes. In previous studies, immobilization of α-amylase on various supports such as magnetic nanoparticles [9], metal ion affinity adsorbent [10] microparticles [11], cellulose membrane [12], nanosphere [13], composite cryogels [14] and beads [15] has been conducted.…”

Immobilization of xylanase enzyme on poly-(HEMA-co-GMA) cryogel

“…Cryogels have 3D elastic interconnected macropores, spongy morphologies and good mechanical robustness [3]. They are extremely versatile tools when converted to desired purposes [4] such as purification/adsorption of biomolecules [5][6][7][8], and removal of some pollutants [9][10][11]. Increasing the existing properties while designing cryogels will further increase their potential use in different fields [4].…”

Investigation of BSA adsorption performances of metal ion attached mineral particles embedded cryogel discs

“…Immobilized metal affinity chromatography (IMAC) presented by Porath et al (1975) has turned into a common analytic and preparative separation technique on purification of some biomolecules [9,10,11,12,13]. Proteins with standard of high purity, which are extensively utilized in various fields, such as molecular biology and biotechnology, can be gained through His-tagged recombinant proteins which require IMAC [14,10].…”

“…They have spongy morphologies with inter-connected macroporous, elastic structures and high mechanical robustness [16]. They are multi-purpose tools that could be turn to good purpose intended practices [17], such as purification and adsorption of some biomolecules [18,19,9], removal of heavy metals, dyes, [20,21] and water treatment process [22]. The boosting of cryogel properties will carry forward its potential usage in various areas [17].…”