Thermodynamic and Kinetic Analysis of Galactose Oxidase Direct Electron Transfer on Carboxyl-Terminated SAM-Modified Gold Electrodes

This study addresses the thermodynamic aspects of galactose oxidase (GAOx) adsorption and redox behavior on gold electrodes modified with self-assembled monolayers (SAMs) derived from thiocarboxylic acids, namely N-acetyl-L-cysteine (NAC), mercaptosuccinic acid (MSA), mercaptoacetic acid (MAA), and L-cysteine (Cys). The electrochemical response of GAOx immobilized on these SAM-modified surfaces was analyzed to extract key thermodynamic parameters governing enzyme-electrode interactions, including the formal redox potential (E°), surface excess (Γ), potential of zero charge (E_zc), adsorption free energy (∆G_add), differential capacitance (C_dl), and surface tension (γ). The results demonstrate that the nature of the terminal functional group of the SAM significantly influences the thermodynamic stabilization of GAOx at the gold interface. Shifts in the redox potential are attributed to specific coordination and electrostatic interactions between the SAM functional groups and the GAOx metal center, leading to distinct interfacial energy landscapes. Overall, the SAM-modified electrodes provide a well-defined thermodynamic framework to probe enzyme orientation, interfacial charge distribution, and stabilization of the redox-active state of GAOx during direct electron transfer. These results offer guidelines based on thermodynamic and kinetic principles for customizing enzyme-electrode interfaces, which can enhance the efficiency, stability, and consistency of third-generation electrochemical biosensors.