Abstract
Novel hybrid magnetic cross-linked enzyme aggregates of phenylalanine ammonia lyase (HM-PAL-CLEAs) were developed by co-aggregation of enzyme aggregates with magnetite nanoparticles and subsequent crosslinking with glutaraldehyde. The HM-PAL-CLEAs can be easily separated from the reaction mixture by using an external magnetic field. Analysis by scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM) indicated that PAL-CLEAs were inlayed in nanoparticle aggregates. The HM-PAL-CLEAs revealed a broader limit in optimal pH compared to free enzyme and PAL-CLEAs. Although there is no big difference in Km of enzyme in CLEAs and HM-PAL-CLEAs, Vmax of HM-PAL-CLEAs is about 1.75 times higher than that of CLEAs. Compared with free enzyme and PAL-CLEAs, the HM-PAL-CLEAs also exhibited the highest thermal stability, denaturant stability and storage stability. The HM-PAL-CLEAs retained 30% initial activity even after 11 cycles of reuse, whereas PAL-CLEAs retained 35% of its initial activity only after 7 cycles. These results indicated that hybrid magnetic CLEAs technology might be used as a feasible and efficient solution for improving properties of immobilized enzyme in industrial application.
Highlights
In the past twenty years, a carrier-free immobilized enzyme strategy, cross-linked enzyme aggregates (CLEAs), has attracted increasing attentions due to its simplicity in preparation and robustness of the immobilized enzymes
The preparation of HM-PAL-CLEAs was elucidated by non-statistical technology, the results revealed that the crude enzyme concentration, magnetite nanoparticles concentration and glutaraldehyde concentration were supposed to have effects on recovery activity
CLEAs technology is attractive in its simplicity and robustness compared with other carrier-free immobilized enzymes technology [2,4,25,31], some undesirable properties limited its further application
Summary
In the past twenty years, a carrier-free immobilized enzyme strategy, cross-linked enzyme aggregates (CLEAs), has attracted increasing attentions due to its simplicity in preparation and robustness of the immobilized enzymes. CLEAs technology has some disadvantages, such as, the particle size of CLEAs is usually small (below 10 mm), which results in difficulties in recovering CLEAs from the reaction medium. They may still be considered too soft for many industrial applications [8,9,10,11]. The supported CLEAs strategies have advantages, the volumetric activity of the biocatalyst and productivity of the reaction can be reduced due to the presence of the noncatalytic mass of the carrier
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