Influence of immobilization protocol on the structure and function of surface bound proteins.

A new coupling strategy for biomacromolecules with (3-mercaptopropyl)trimethoxysilane (3MPTMS) and 11-(triethoxysilyl)undecanal (TESU) on gold surfaces is. This immobilization protocol was utilized for the enzyme horseradish peroxidase (HRP). To study the reactions and resulting structures, PM-IRRAS measurements were performed. PM-IRRAS shows there is structure preservation of the HRP when the new coupling strategy is used in contrast to non-specific adsorption on gold. The biological activity of adsorbed and immobilized HRP was measured by the enzyme catalyzed oxidation of 3,5,3',5'-tetramethylbenzidine. Covalent immobilization of HRP on TESU film compared to physisorption of HRP shows higher enzyme activity on gold surfaces, confirming the structural preservation detected by PM-IRRAS.