A highly selective protein adsorber via two-step surface-initiated molecular imprinting utilizing a multi-functional polymeric scaffold on a macroporous cellulose membrane

Abstract

Although molecularly imprinted materials using small organic molecules as templates have been well established, development of such materials for protein separation is still rather challenging. We therefore describe herein a two-step surface imprinting method established with a hydrophilic macroporous cellulose membrane with relatively large specific surface area. In the first step, tailor-made multi-functional polymer chains were grafted on the cellulose membrane using photo-initiated graft copolymerization, enabled by a surface-immobilized photo-initiator. This scaffold allowed the preorganization of the template protein lysozyme (Lys) on the surface of the membrane pores. Notably, the scaffold-grafted membrane showed already a significant adsorption selectivity versus the very similar protein cytochrome C (CyC). In the second step, surface-initiated cross-linking copolymerization, enabled by a photo-initiator immobilized in the scaffold layer, resulted in a protein-imprinted cellulose membrane. Imprinting efficiency was further improved by optimization of monomer concentrations in the second step. Protein selectivity of the best imprinted cellulose membrane for Lys over CyC reached a very remarkable value of about 45, measured with 1 : 1 mixtures of the two proteins. We envision that this property of a protein-imprinted cellulose membrane, which is based on the tailored binding selectivity achieved using the two-step functionalization method, could be largely beneficial for separation and purification of target proteins from complex mixtures.