Site directed confinement of laccases in a porous scaffold towards robustness and selectivity

• Published in: Biotechnology reports (Amsterdam, Netherlands) Netherlands), 2021-09, Vol.31, p.e00645-e00645, Article e00645
• Main Authors: Yang, Fangfang, Backov, Rénal, Blin, Jean-Luc, Fáklya, Bernadett, Tron, Thierry, Mekmouche, Yasmina
• Format: Article
• Language: English
• Subjects: Biotechnology; Heterogeneous catalysis; Laccase; Life Sciences; Orientation; Site-directed immobilization
• ISSN: 2215-017X; 2215-017X
• DOI: 10.1016/j.btre.2021.e00645

[Display omitted] •Site directed immobilization of laccase on silica foam via covalent bond forming reaction.•RB5 decolorization activity is maintained for 13 reaction cycles or one year storage towards RB5 oxidation.•Active site orientation towards porous material decreases/increases large substrate oxidation. We immobilized a fungal laccase with only two spatially close lysines available for functionalization into macrocellular Si(HIPE) monoliths for the purpose of continuous flow catalysis. Immobilization (30–45 % protein immobilization yields) was obtained using a covalent bond forming reaction between the enzyme and low glutaraldehyde (0.625 % (w/w)) functionalized foams. Testing primarily HBT-mediated RB5 dye decolorization in continuous flow reactors, we show that the activity of the heterogeneous catalyst is comparable to its homogeneous counterpart. More, its operational activity remains as high as 60 % after twelve consecutive decolorization cycles as well as after one-year storage, performances remarkable for such a material. We further immobilized two variants of the laccase containing a unique lysine: one located in the vicinity of the substrate oxidation site (K157) and one at the opposite side of this oxidation site (K71) to study the effect of the proximity of the Si(HIPE) surface on enzyme activity. Comparing activities on different substrates for monoliths with differentially oriented catalysts, we show a twofold discrimination for ABTS relative to ascorbate. This study provides ground for the development of neo-functionalized materials that beyond allowing stability and reusability will become synergic partners in the catalytic process.