Smart peptides: immobilization of hyperthermophilic enzymes on mineral matrices for biomass degradation

Biofunctionalization describes the immobilization of biomolecules onto the surfaces of solid matrices to endow them with a biological function.  Most methods available for immobilizing proteins onto solid supports traditionally have relied on nonspecific adsorption or on the reaction of naturally occurring chemical groups such as amines and carboxylic acids within proteins with appropriate reactive groups on the matrix.  In both cases, the corresponding proteins are attached to the surface in a random orientation that may cause the reduction or loss of the protein’s biological activity.

We have developed a versatile biofunctionalization method based on peptide linkers (Smart Peptides) that can direct the immobilization of proteins onto solid surfaces without impeding protein function. This new approach is simple, straight-forward and bypasses conventional techniques such as adsorption, covalent binding, cross-linking and entrapment. The peptide linker sequence displays high affinity towards materials that contain silica. Thus, the linker, as part of a fusion protein in conjunction with materials containing silica, can be used for the isolation, purification or immobilization of the protein. The method is robust and capable of introduction into industrial processes.

Following this approach, we have successfully incorporated the Smart Peptide linker into more than 25 different proteins including industrially-relevant ones such as hyperthermophilic β-1,4-xylanase, β-1,4-mannanase, β-glucosidase and an endo-glucanase and have demonstrated their effective immobilization onto zeolite and silica. We showed the reuse of the immobilized enzymes at high temperatures in multiple sequential enzyme reactions without loss of activity, thus potentially enabling maximization of their economic value.