text: Diisopropyl fluorophosphate (DIFP) is an irreversible inhibitor that blocks the action of serine protease. When it binds to the enzyme a nucleophilic substitution reaction occurs and releases one hydrogen fluoride molecule. The OH group in the active site acts as a nucleophile to attack the phosphorus in DIFP and form a tetrahedral intermediate and release a proton. Then the P-F bond is broken, one electron is transferred to the F atom and it leaves the intermediate as F anion. It combines with a proton in solution to form one HF molecule. A covalent bond formed between the active site and DIFP, so the serine side chain is no longer available to the substrate.