The GFP Rainbow: An Educational Tool for Teaching Molecular Modeling and Molecular Dynamics

This work presents a computational biochemistry activity designed for advanced undergraduate or graduate students who wish to explore the relationship between structure, dynamics, and energetics in Green Fluorescent Protein (GFP) variants through structural biology and computational modeling. The activity integrates practical skills in molecular visualization with conceptual understanding of molecular mechanics, thermodynamics and intermolecular interactions. The students learn to prepare complex systems containing nonstandard residues, such as the GFP fluorophore, generate the corresponding parameter files, and perform GPU-accelerated molecular dynamics simulations under realistic solvated conditions. The resulting trajectories are analyzed to evaluate thermodynamic stability, structural evolution, and energy contributions. Monitoring the evolution of kinetic, potential, and total energies reinforces fundamental principles such as energy conservation and equilibrium. Structural analyses, including Root-Mean-Square Deviation (RMSD), Root-Mean-Square Fluctuation (RMSF) and hydrogen bonding networks, provide insight into protein stability and fluorophore interactions, while energy decomposition quantifies per-residue contributions to fluorophore stabilization. Through this integrated workflow, students connect theoretical principles with computational practice, learning to interpret numerical data in terms of physical meaning. The complete set of input files, scripts, simulation results and analysis for several GFP variants is provided through an open GitHub repository, thus facilitating adoption and adaptation for classroom or research-based learning environments.