Protein imprinting in hydrogels is one approach for developing artificial receptors capable of specific recognition and binding of a target molecule. Through selection of monomers with side groups that can interact with the target protein and control over the degree of cross-linking, the architecture and spatial distribution of interaction points can be optimized for a target protein. Here we report on the imprinting of polyacrylarnide-based hydrogels with maltose binding protein (MBP). To design the optimum architecture, we analyze the distribution of surface amino acid residues on the protein surface. We show that the selectivity of MBP recognition is increased by incorporating monomers that can introduce sites for hydrogen bonding, hydrophilic interactions, and electrostatic interactions. MBP-imprinted films showed high specificity and could discriminate between reference proteins with similar molecular weight, dimensions, and isoelectric point.