p ‐Benzoyl‐ l ‐phenylalanine as a Multifunctional Noncanonical Amino Acid in Synthetic Biology: Photoprobing, Photocatalysis, and Structural Programming for Biocontainment

ABSTRACT p ‐Benzoyl‐ l ‐phenylalanine (pBzF) is a widely used noncanonical amino acid (ncAA) that expands the chemical repertoire of proteins. Its benzophenone (BP) chromophore undergoes near‐quantitative intersystem crossing (ISC) to a triplet state, furnishing a highly efficient, site‐addressable photoreactive handle. Beyond photochemistry, the bulky, hydrophobic side chain introduces distinct steric and electronic effects that enable new reactivity in protein active sites. Genetic incorporation of pBzF in vivo, including directed evolution, has unlocked applications ranging from site‐specific photo‐crosslinking for interaction mapping to engineering antibody fragments, sharpening monoclonal antibody (mAb) epitope recognition, and creating protein‐based photocatalysts. pBzF has also proved powerful for mechanistic studies by stabilizing short‐lived intermediates. More recently, pBzF‐containing proteins have been leveraged in light‐driven transformations, including 2+2 photocycloadditions, deracemizations, and dehalogenations, and in the construction of artificial photosynthetic systems. This review critically discusses these advances and establishes pBzF as a versatile photochemical and structural motif for building proteins with non‐natural, light‐responsive, and catalytically competent functions.