N-Formylglycine is widely used in biochemical research, particularly in studies of sulfatase enzyme mechanisms. Sulfatases contain a post-translationally generated formylglycine (FGly) residue—a structural isomer of N-Formylglycine—that acts as a key intermediate in their catalytic cycle for hydrolyzing sulfate esters. N-Formylglycine serves as a valuable model compound, synthetic precursor, or probe to investigate and manipulate sulfatase activity, which is essential for biological processes such as hormone metabolism and complex carbohydrate degradation. It is also applied in developing enzyme inhibitors, designing therapeutics for sulfatase-related diseases, and synthetic biology for engineering enzymes with improved functions for industrial and medical applications.