In the present work the interaction between GST (glutathione-S-transferase) and immobilized glutathione on the mixed thiol monolayers was investigated by surface plasmon resonance method. The interfacial structures based on mixed monolayer of CH3-/C(O)OH and OH-/C(O)OH- terminated thiols as an support for the immobilization of glutathione (GSH)where used. The range of matrix/reactive groups ratio was up to 1:700. The standard procedure with activation by NHS/EDC with further applying of PDEA was used for GSH binding. It was demonstrated, that both the nature of the matrix terminal group and the density of the immobilized ligand are important for the efficiency of the protein-surface and protein-protein interaction at the sensor surface. Testing of the surface coupled GST by specific anti-GST antibodies shown the same character of the sensor response versus GSH density.
Using surface plasmon resonance we studied how the ferment- substrate interaction between trypsin and its soybean inhibitor at gold surface depended on previous treatment of the gold surface with NaNCS (thiocyanate) and NaNCO. It was shown that protein adsorption appreciably increased after surface modification by thiocyanate and didn't show any increase after their treatment with NaNCO. In the case of NaNCS treatment protein retained its biological properties and formed an oriented monolayer.
The formation of Au<SUB>x</SUB>S<SUB>y</SUB> interfacial layer by reactive annealing of gold films in H<SUB>2</SUB>S atmosphere is investigated. This seems to be a technologically favorable technique for the large-scale and low-cost fabrication of nondestructive immobilization support for biological molecules. Formation of phases with different chemical functionality and surface topography as a function of reaction time was studied using Atomic Force Microscopy (AFM), Surface Plasmon Resonance (SPR) measurements and biomolecular interaction analysis [trypsin -- Soybean Trypsin Inhibitor (STI) reaction]. The results obtained confirm the classical two-step model for the sulfide phase formation during reactive annealing. This includes an intermediate formation of a dispersed phase of sulfur followed by its reconstruction to a close-packed sulfide layer. Adsorption of proteins onto a certain sulfide layer passes with retention of a native state of adsorbed molecules. The proposed strategy for formation of biochemical structures (gold/sulfide/proteins) on the surface of physical transducers open a new way for design and development of novel artificial smart sensing systems. They not only maintain optimal functioning of bioreceptors but also are responsive to their environment.