The coxibs are a subset of non-steroidal anti-inflammatory drugs (NSAIDs) that selectively target cyclooxygenase-2 (COX-2) to inhibit prostaglandin signaling and reduce inflammation. However, only celecoxib remains in use, necessitating exploration of broader mechanisms of the coxibs. Here, we report a novel binding site for celecoxib on prostaglandin E synthase (PTGES), an enzyme downstream of COX-2 in the prostaglandin signaling pathway using an isotopically-coded cleavable chelation-assisted biotin probe. Evaluation of the multi-functional probe revealed significantly improved tagging efficiencies attributable to promotion of CuAAC chemistry by the embedded picolyl functional group. Application of the probe within the small molecule interactome mapping by photo-affinity labeling (SIM-PAL) platform using photo-celecoxib as a reporter for celecoxib identified known targets (e.g., carbonic anhydrase 12) and the significant enrichment of PTGES, along with six additional membrane proteins and 15 subunits of the cytochrome complex. In addition, four binding sites to celecoxib were mapped by the probe, including a direct interaction with PTGES. The interaction between celecoxib and PTGES was validated by competitive displacement and thermal shift assay. The binding site between celecoxib and PTGES enabled the development of a structural model of the interaction and will inform the further development of new selective inhibitors of the prostaglandin signaling pathway.