Thermophilic esterase from the archaeon Archaeoglobus fulgidus physically immobilized on hydrophobic macroporous resin: A novel biocatalyst for polyester synthesis

Abstract

This paper reviews the immobilization of a thermophilic esterase, AFEST from the archaeon Archaeoglobus fulgidus, on a hydrophobic macroporous resin and its application in polyester synthesis using the ring-opening polymerization of ε-caprolactone as a model. Using the physical adsorption technique, the AFEST loading concentration after 24 h was 152 mg AFEST per g of support.Particle size and surface morphology of the immobilized enzyme were investigated using laser scattering analysis and scanning electron microscopy. The effects of enzyme concentration, temperature, reaction time and reaction medium on monomer conversion and product molecular weight were systematically investigated. Through the optimization of reaction parameters, poly(ε-caprolactone) was obtained at an almost 100% monomer conversion rate and with a low average molecular weight (< 1,100 g/mol). Finally, the immobilized enzyme exhibited good operational stability, with a monomer conversion value of more than 55% after four batch reactions.

Glutaraldehyde Cross-Linking of Immobilized Thermophilic Esterase on Hydrophobic Macroporous Resin for Application in Poly(ε-caprolactone) Synthesis

“…It has been generally accepted that the reaction medium plays a crucial role in determining the stability of the biocatalyst and in the partitioning of substrates and products between the solvent and the biocatalyst in non-aqueous biocatalytic systems [32]. Table 1 showed the effect of organic solvents with different Log P values on monomer conversion and M n at 70 °C for 72 h. Similar to our previous reports [13,14,24], hydrophobic solvents (toluene, cyclohexane and n -hexane) were favorable for the reaction, with high monomer conversion (100%) and M n values (>950 g/mol). The phenomenon was caused by the fact that the hydrophobic solvents could efficiently keep the essential water layer surrounding the enzyme molecules and did not disrupt the functional structure of enzymes [33].…”

“…As reported previously [13], the thermophilic esterase AFEST was partially purified, with an active protein content of 92% (w/w) in the enzyme preparation, and the specific activity of lyophilized enzyme was approximately 168 U/mg toward p -nitrophenyl caprylate at 80 °C. Through the physical adsorption for 24 h, the adsorption efficiency was measured to be 76%, and thus the loading of AFEST on supports was calculated as 152 mg AFEST/g supports [24]. After the physical adsorption, cross‑linking of enzymes on the surface of supports was then conducted through the treatment with different concentrations of glutaraldehyde.…”

“…As the support is highly hydrophobic, it could adsorb the open form of lipase (fixed on the interfacially adsorbed derivative), and the immobilized lipase exhibited higher activity and stability [25]. However, in our research, the immobilized enzyme possessed only 36% of the original synthetic activity after five reuses, probably due to the loss of adsorbed enzyme from the support during consecutive reactions [24,26]. In fact, all these immobilized enzymes are used with the problem of limited operational stability, and thus developing more effective biocatalysts through chemical crosslinkages or protein modification will be of great significance for realizing enzymatic polymerization at an industrial scale in future.…”

“…50,000 g/mol [23]. Similarly, our research group adsorbed AFEST to the hydrophobic macroporous resin Octadecyl-Sepabeads EC-OD and used it for the synthesis of PCL [24]. As the support is highly hydrophobic, it could adsorb the open form of lipase (fixed on the interfacially adsorbed derivative), and the immobilized lipase exhibited higher activity and stability [25].…”

A Review on Extremozymes Biocatalysis: A Green Industrial Approach for Biomaterials Production

“…Moreover, compared with the reported lipases, AFEST had the highest affinity for ε-caprolactone as demonstrated by Michaelis -Menten kinetic analysis and molecular docking. AFEST was improved by G. Li and Q. Li by immobilization on hydrophobic macroporous resin [76]. The use of immobilized enzymes not only can have a good effect on the enzyme stability, can produce economic benefits to the biocatalytic process through enzyme reuse, but also can be a method of election to scale processes from laboratory to industry.…”