Highly selective oligosaccharide production through integration of enzymatic synthesis and peptide affinity under non-conventional process conditions

For many biotechnological processes, a large part of the manufacturing costs and a large part of the use of resources is needed for product recovery and purification. Further improvement will need the development of integrated production systems, combining synthesis and purification. In this proposal, we aim for the development of a generic process, combing the use of biocatalysts and the use of the biomolecular recognition. By using selective biocomplexants, with an aspecific biocatalyst, a highly selective process can be developed. Because of the wide diversity of biocomplexants found in nature, the process can be made very flexible. In this proposal we focus on the production of specific oligosaccharides. Oligosaccharides are receiving much attention as biofunctional components, for nutritional purposes, but also their pharmaceutical application gains more and more interest. The enzymatic synthesis using mono- and/or disaccharides as substrates results in the production of a wide range of oligosaccharides, of diverse chain length, monomer sequence and chain morphology. This could be improved by introducing a highly specific, in-line isolation of the product. In nature, several peptides are known that show a high complexation affinity towards specific oligosaccharides. In this project, we will use the specific binding capacity of lectins and the so-called Designer Affinity Molecule (DAM), developed by CatchMabs. The proteineous designer affinity molecules are extremely stable and the molecular structure is very flexible and quickly adaptable. The concept proposed is the combination of an aspecific enzymatic reaction with a more specific process for product removal. Since the affinity of certain peptides with specific oligosaccharides is very high, it is possible to apply the system at very high concentrations. This is thermodynamically favorable, and can lead to high productivities. Especially interesting is operation at higher temperatures (50-90ºC), since this increases the maximum substrate concentration considerably and can therefore lead to a significant increase in productivity. The enzyme is freely suspended in the reaction mixture, the peptides are immobilized. The desired product can be effectively removed e.g. by using magnetic particles for immobilization of peptides. The magnetic particles can easily removed from the reaction mixture. Attention will be paid to the regeneration of the immobilized particles since the binding is not only selective, but also strong. The project will involve selection of suitable peptides (lectins as well as DAMs), immobilization of peptides on magnetic particles, development of an effective and efficient downstream process (desorption and final purification) to come to oligosaccharides with a high purity and development of an effective integrated process design, in which flexibility towards specific oligosaccharides can be as high as possible. In this project we apply for two AIO’s, one at Organic Chemistry, focusing on screening and selection of peptides and one at Process Engineering, focusing on downstream processing and process design.