Pulsed-laser metal contacting of biosensors on the basis of crystalline enzyme-protein layer composites

Abstract

Crystalline bacterial cell surface layers (S-layers) composed of monomolecular arrays of protein subunits are accessible to a wide variety of possible proteinchemical reactions. This enables the attachment and immobilization of enzyme molecules in a tightest packing, which has not been achieved with other immobilization matrices. When immobilized to an S-layer lattice, the enzyme entities are surrounded by nanometer pores. Thus, they can react electrochemically with the analyte liquid streaming through these pores. The control over this process has to take place by way of an inert electrical contact in a distance of less than 1 nm. The relatively voluminous, but specially shaped sensor enzyme molecules have to be connected with an optimum metallic contact, which must not disturb the protein structure. Previously, platinum films were applied on enzyme layers immobilized on S-layer protein by argon sputtering. This conventional technique, however, exhibits substantial limitations. One, for instance, is the volume change of the S-layer/enzyme composite system when it is introduced into a conventional vacuum coating apparatus. This coating problem can be circumvented by a completely new deposition method, i.e. the pulse-laser-deposition (PLD) on protein crystal composite films with optimized laser parameters and reaction atmospheres. Enzyme activities of 70–80% were achieved, thus demonstrating that composite systems consisting of the 2D-protein-layer/enzyme/metal sequence can successfully serve as highly efficient sensor systems.