Prediction of residues involved in inhibitor specificity in the dihydrofolate reductase family

► Methods were developed to quantify similarity in inhibitor specificity for homologs. ► Homology-based process was developed to predict inhibitor specificity determinants. ► Eighteen DHFR positions that play a role in ligand specificity were identified. ► For these, amino acid sequence homology correlates with ligand specificity. ► Three are in active site, 5 near the active site, the rest may act allosterically.