Electrochemical and Binding Studies of Solvent Reorganization Energy ProbesPublic Deposited
In this study, ligands modified to contain a metal ion were synthesized and their electrochemical properties were examined alone and in the presence of a protein. We chose to use avidin and biotin as a model system. Probes have been developed in which biotin is modified to coordinate to a metal, specifically ruthenium. Binding to avidin displaces solvent molecules and changes the optical and static dielectric constants in the outer sphere environment of the metal ion. The electrochemistry of these complexes was investigated in the presence and absence of avidin. The expected shift in peak potential of the ruthenium complexes was not observed; rather, a lack of current signal in the cyclic voltammogram resulted in the presence of avidin. Electrochemical experiments using mediators were conducted. The mediator, (4-chloropyridine)Ru(NH3 ) 5 3+, and binding complex, (4-DMP)Ru(NH3 ) 5 3+, were synthesized and characterized via UV-visible spectroscopy. Binding between (4-DMP)Ru(NH3 ) 5 3+ and avidin was characterized using HABA assays, which show that the 4-DMP complex binds to avidin in a ratio of four moles of binding complex to one mole of protein, as expected. The dissociation constant for this binding complex was determined to be 1.8 × 10-8 M. Conditions have been determined for the handling of the binding complex and the electrochemical experiments that yield data suitable for simulation fitting. Cyclic voltammetry experiments were conducted at multiple concentrations, and scan rates and simulations are under way to determine the rate of electron transfer.