Small ubiquitin-like modifier (SUMO)-activating enzyme (E1) has been shown previously as a target to eradiate cancer stem cells of various cancer types. In this study, we discovered a novel mechanism to inhibit this target through a previously unknown site of the enzyme resulting in a major change of the shape of the enzyme to destroy its catalytic activity. Targeting this site not only destroys the enzyme activity of the SUMO E1, but also enhances its degradation in vivo, presumably due to a conformational change induced by the compound. The mechanism we identified is applicable to other family members of this class of enzymes and will spur new innovation in therapeutic discovery targeting new signaling pathways that cancer stem cell relies on.