Bio-inspired enzyme entrapment and cross-linking approaches as alternative tools for enzyme immobilization

Ardao Palacios, Inés [UCL] Demarche, Philippe [UCL] Nair, Rakesh [UCL] Agathos, Spiros N. [UCL]

Enzyme immobilization has contributed to the widespread use of enzymes as catalysts in many industrial applications mainly due to stability enhancement and easy reuse of the catalysts, which contributes to a reduction of the process costs. However, enzyme immobilization still suffers from different operational constraints, such as activity losses during immobilization and mass-transfer limitations. Novel immobilization techniques with the goal of overcoming these limitations are increasingly developed, one of the most promising being the immobilization on nanoparticles and nanostructures Recently, bio-inspired enzyme entrapment by biomimetic mineralization has emerged as a versatile tool for generating robust, cheap and highly stable (nano)biocatalysts. In analogy to the natural silica precipitating peptide templates (silaffins) of diatoms, other polymer templates (either natural or synthetic analogues) can be used to induce the precipitation of different inorganic oxides resulting in the entrapment of enzymes and other materials present in the medium. Different (nano)biocatalysts have been produced in our group following different immobilization approaches: i) cross-linking aggregates of single enzymes (CLEAs) or enzyme cascades (Combi-CLEAs); ii) combination of enzyme adsorption and cross-linking on functionalized mesoporous silica particles and nanoparticles; and iii) bio-inspired enzyme entrapment by biomineralization. Oxidative enzymes (fungal laccases, peroxidases and oxidases) were used as model enzymes. These enzymes are able to catalyze oxidation reactions using molecular oxygen or hydrogen peroxide as co-substrates and are, therefore, highly interesting for biocatalytic and environmental applications. The produced (nano)biocatalysts have been successfully applied to the degradation of micropollutants (endocrine disruptor chemicals), showing a high degradation capacity in a continuous reactor for the treatment of real secondary effluents from a wastewater treatment plant. The high versatility of the biocatalysts and enzyme immobilization techniques presented here allows envisaging promising applications on the development of new bioproduction systems based on immobilization of natural or synthetic biocatalysts and enzyme cascades.