Aminated laccases can improve and expand the immobilization protocol on agarose-based supports by ion exchange

Abstract

In this work, laccases were immobilized onto positively charged supports such as Monoaminoethyl-N-aminoethyl (MANAE)-agarose and Diethylamino ethyl (DEAE)-agarose and onto negatively charged support (sulfopropyl agarose). However, the latter required prior enrichment of the protein surface with amino groups. The amination process was carefully controlled by varying concentrations of 1-ethyl-3-(dimethylamino-propyl) carbodiimide (EDAC), resulting in three types of aminated laccase: SA1 (3.364 mM of EDAC), SA2 (10.09 mM of EDAC), and SA3 (23.55 mM of EDAC). According to the CD spectra, the introduction of amino groups on the enzyme surface caused conformational changes that either positively (SA1) or negatively (SA2 and SA3) impacted the secondary structure, depending on the concentration of EDAC used. In addition, the concentration of EDAC during amination positively affected the immobilization yield on the sulfopropyl support. Amination also facilitated the production of biocatalysts with enhanced stability, with half-lives of 78 and 90 min for SA2 and SA3 preparations at 60 °C and pH 7.0. These findings imply that aminating laccase prior to immobilization is a valuable approach for broadening the versatility and usefulness of laccase-based systems. As a result, heterogeneous biocatalysts with improved properties have been prepared, expanding the range of potential applications.