Study of silanized nanostructures with immobilized fumarase for production of L-malate

Abstract

The conversion of fumaric acid into L-malate by fumarase immobilized on silanized nanostructures was analyzed experimentally. The enzyme was bound to the silanized nanostructures. We carried out scanning electron microscopy (SEM), fourier transform infrared spectroscopy (FTIR) analysis, zeta size analysis and surface area calculation for the characterization of the nanostructures. The effect of initial enzyme concentration and pH on immobilization procedure were investigated and the change of Michaelis-Menten constants (K m and V max ) with immobilization was examined. The change in the storage stability of the enzyme by immobilization was also investigated. The stability of the immobilized enzyme was very good. We observed that the fumarase was bound to silanized nanostructures [p(HEMA)-3-MTES] in much greater amounts. We have compared the activities of free fumarase and immobilized fumarase and we have observed a significant increase in the activity of the fumarase after immobilization for L-malate production. Moreover, we came to the conclusion that this activity can be better preserved for 30 days compared to free fumarase.

Immobilization of fumarase from thermophilic eukaryotic red alga <i>Cyanidioschyzon merolae</i> on ceramic carrier

“…On the other hand, Atay et al (2022) reported a slight decrease in the Km value from 5.55 mM to 5.05 mM and an approximately 1.8-fold increase in reaction velocity when immobilizing pig heart fumarase on poly(hydroxyethyl methacrylate) silanized with 3-mercaptopropyltriethoxysilane. In their approach, enzymes are immobilized on the carrier through a disulfide bond between the surface-positioned cysteine residues of the enzyme and the thiol groups of (3-mercaptopropyl) triethoxysilane.…”

“…The binding of hydroxy groups on the carrier to areas close to the substrate binding site might interfere with access or binding of substrate to the substrate binding site, thereby leading to an increased Km value. However, although an increase in Km value was observed after immobilization, the lower Km value of immobilized CmFUM for fumarate compared to other immobilized fumarases suggests a superior affinity of immobilized CmFUM for fumarate (Atay et al, 2022;Giorno et al, 2001;Li et al, 2022;Marconi et al, 2001, Table 3). On the other hand, decrease in kcat was significant and was thought to be a major factor associated with the decrease in CmFUM activity following immobilization.…”

“…Recently, Li et al (2022) reported that fumarase from Thermus thermophilus immobilized on konjac-κ-carrageenan beads showed improved stability during storage and high reusability, although its enzymatic activity decreased (Li et al, 2022). Atay et al (2022) have reported that immobilized fumarase from pig heart on silanized nanostructures shows improved storage stability and increased activity (Atay et al, 2022). Immobilized enzymes are expected to be exposed to reaction heat for extended periods, but to date there have been no detailed reports of the thermostability of immobilized fumarase.…”