Immobilization of enzymes onto solid supports can be achieved by a number of physical and chemical methods including, covalent attachment, adsorption, crosslinking and encapsulation. Immobilized enzymes, as opposed to soluble enzymes, offer better stability and easier removal from reaction mixtures, enabling repetitive use in batch and continuous bioprocesses and rapid termination of reactions.
Unfortunately, typical enzyme immobilization approaches usually result in a non-uniform orientation of the enzyme as well as unwanted conformational changes that alter their active sites and may curtail the catalytic activity of the enzyme. Solid-binding peptides have binding affinity as well as selectivity to the surfaces of solid materials such as glass, polymers, silica, metals and zeolite, all support materials employed with biocatalysts.
Solid-binding peptides typically are used as molecular linkers for functional protein immobilization onto solid surfaces without the need for any chemical reactions or even physical treatments.
Researchers led by Professor Anwar Sunna at Macquarie University in Australia have presented the implementation of the solid-binding peptide-mediated immobilization of industrially-based enzymes onto a low cost solid zeolite matrix. The introduction of crosslinking of the immobilized enzymes to create single as well as multiple enzyme biocatalytic modules enabled the authors to highlight the feasibility of this technology for its integration in industrial-scale processes. Their work is published in Biotechnology for Biofuels.
The research team genetically fused the silica-binding linker peptide to three thermostable polysaccharide-degrading enzymes for potential application in industrial-scale biocatalysis. The linker had significant affinity for silica-containing supports allowing for directional immobilization of these enzymes onto the zeolite matrix. The enzymes were observed to retain their binding affinity for zeolite and their biological activity. The integration of the linker did not have adverse effects on the pH and temperature optima of the polysaccharide-degrading enzymes and the assembled single and multiple enzyme biocatalytic modules retained their specific hydrolytic activities upon several rounds of recycling at high temperatures.
Professor Sunna summarized the importance of this platform technology saying; “Despite the promising characteristics of solid-binding peptides, their practical application has been mostly in nanobiotechnology, where immobilization of biomolecules generally relies on exotic and expensive laboratory-based matrices that may not be realistic economically for large-scale processes. Inorganic bulk materials like zeolite and silica are excellent carriers due to their structural and operational stability and their lack of susceptibility to microbial degradation. The combination of solid-binding ability and low-cost bulk materials represents an ideal technology for production of industrial-scale biocatalysts.”
The linker system developed in their study minimizes the time wasted in choosing precipitants as well as crosslinking reagents. Its compositional and structural characteristics allows it to impart orientation and directionality to enzymes after crosslinking. This combination results in improved enzyme reusability. Therefore, this linker technology presents an inexpensive immobilization approach for industrially based enzymes.
Andrew Care, Kerstin Petroll, Emily S. Y. Gibson, Peter L. Bergquist, and Anwar Sunna. Solid-binding peptides for immobilization of thermostable enzymes to hydrolyze biomass polysaccharides. Biotechnol Biofuels (2017) 10:29.Go To Biotechnology for Biofuels