D-4-Cyanophenylalanine is primarily used in biochemical research as a non-natural amino acid. It serves as a valuable tool in protein engineering and structural studies, particularly in fluorescence spectroscopy and Förster resonance energy transfer (FRET) experiments. Its unique cyano group allows it to act as a sensitive probe for monitoring protein folding, conformational changes, and interactions within biological systems. Additionally, it is employed in the development of biosensors and in studying enzyme mechanisms, where its incorporation into peptides or proteins provides insights into molecular dynamics and function. Its applications also extend to drug discovery, where it aids in the design and optimization of therapeutic compounds.