Integrating of Common Mechanisms in Protein-Film Voltammetry-MATHCAD Simulation Protocol

Protein-film voltammetry (PFV) provides a powerful methodology for investigating electron-transfer processes and coupled chemical reactions of surface-confined proteins and enzymes. In this work, for the firt time, a comprehensive theoretical framework is presented for the surface CirrE₁CrevE₂C′ mechanism, integrating a preceding irreversible chemical transformation, two consecutive electron-transfer steps interconnected by a reversible chemical reaction, and a final regenerative step. Owing to its multistep architecture, the model encompasses several commonly encountered PFV mechanisms as limiting cases, providing a unified platform for their systematic analysis. Simulated cyclic voltammetric patterns demonstrate how the kinetics of individual chemical and electron-transfer steps, the equilibrium constant of the intermediate reversible reaction, and catalytic regeneration govern the shape, magnitude, reversibility, and potential position of the voltammetric responses. Particular attention is given to systems with closely spaced electron-transfer potentials, where catalytic kinetics can assist in resolving otherwise overlapping redox processes. Importantly, the original Mathcad simulation protocol for the surface CirrE₁CrevE₂C′ mechanism in cyclic voltammetry is provided, enabling straightforward variation of relevant kinetic and thermodynamic parameters. The presented framework offers a practical computational platform for simulation, mechanistic exploration, and interpretation of complex reaction pathways encountered in protein-film voltammetry.