Selective metal ion affinity binding as a simple and renewable enzyme immobilization was investigated using the same affinity function on various supports. Phenylalanine ammonia-lyase from parsley (PcPAL) and an amine transaminase from Vibrio fluvialis (VfTA) with His-tag were used as model enzymes. Metal ion chelating groups on the surface of six enzyme carriers were created from the surface-alkylamino moieties by treatment with ethylenediaminetetraacetic dianhydride and subsequent complexation with cobalt(II) ions. Three porous polymer beads, two silica-based supports, and a silica-coated magnetic nanoparticle (MNP) were investigated as carriers. The most effective PcPAL biocatalyst forms were tested in kinetic resolution and ammonia addition reactions with substrates containing phenyl and thiophen-2-yl rings. In the selective ammonia addition reaction onto the (hetero)arylacrylates needing a harsh medium of a 6 M ammonia solution, the biocatalysts exhibited excellent stability and led to l-amino acids in high yield and excellent enantiomeric excess. The recharging of the MNP supports was investigated by five subsequent cycles of reactions-after elution with 5% diethylenetriamine and reloading with fresh PcPAL or VfTA-retaining over 80% of the relative activities until the third cycle. The repurposing of the supports was also investigated by changing one enzyme to the other on the MNP-immobilized metal ion affinity chromatography support.
Laurinyecz et al. (Wed,) studied this question.