It is well established that various endocrine disrupting compounds (EDCs) can inhibit human estrogen sulfotransferase (SULT1E1). In this study, we investigate murine SULT1E1 inhibition in vitro and in silico and compare this to data for the human enzyme. 34 potential EDCs were screened for their ability to inhibit both murine and human SULT1E1 and IC(50) values were determined for 14 of the inhibitory EDCs. Only estrone, dienestrol and enterolactone showed significant differences in affinity between the human and murine SULT1E1. Extensive molecular modelling was performed using molecular dynamics (MD) simulations. During the MD simulations the ligands moved away from the catalytically active position, something which was not observed when simulating the unit cell of the crystal structure. This finding suggests that catalytically inactive binding modes, other than the one observed in the crystal structures, are possible in SULT1E1. The ligands stayed longer in the catalytically active position in mSULT1E1, which is likely a result of simultaneous hydrogen bond formation on both sides of the binding pocket, which does not seem to be possible in hSULT1E1. The ligands in the human protein moved to a sub-pocket near the entrance of the active site, which offers hydrogen bond formation possibilities with Asp22 and Lys85 as well as favourable hydrophobic interactions. The ligands moved more randomly in mSULT1E1. These observations offer a possible explanation for the substrate inhibition only observed in hSULT1E1.