It is increasingly clear that interindividual variability in human gut microbial composition contributes to differential drug responses. For example, gastrointestinal (GI) toxicity is not observed in all patients treated with the anticancer drug
irinotecan, and it has been suggested that this variability is a result of differences in the types and levels of gut bacterial β-glucuronidases (GUSs). GUS
enzymes promote
drug toxicity by hydrolyzing the inactive drug-
glucuronide conjugate back to the active drug, which damages the GI epithelium. Proteomics-based identification of the exact GUS
enzymes responsible for drug reactivation from the complexity of the human microbiota has not been accomplished, however. Here, we discover the specific bacterial GUS
enzymes that generate
SN-38, the active and toxic metabolite of
irinotecan, from human fecal samples using a unique activity-based
protein profiling (ABPP) platform. We identify and quantify gut bacterial GUS
enzymes from human feces with an ABPP-enabled proteomics pipeline and then integrate this information with ex vivo kinetics to pinpoint the specific GUS
enzymes responsible for
SN-38 reactivation. Furthermore, the same approach also reveals the molecular basis for differential gut bacterial GUS inhibition observed between human fecal samples. Taken together, this work provides an unprecedented technical and bioinformatics pipeline to discover the microbial
enzymes responsible for specific reactions from the complexity of human feces. Identifying such microbial
enzymes may lead to precision
biomarkers and novel drug targets to advance the promise of
personalized medicine.