Human estrogen receptor alpha (ERα), which acts as a biomarker and as a therapeutic target for breast cancers, is activated by agonist ligands and co-activator proteins. Selective estrogen receptor modulators (SERM) act as antagonists in specific tissues and tamoxifen, a SERM, has served as a drug for decades for ERα-positive breast cancers. However, the ligand-selective and tissue-specific response of ERα biological activity and the resistance to tamoxifen treatment in advanced stages of ERα-positive breast cancers underscores the need to find a ligand-independent inhibitor for ERα. Here we present a ligand-independent approach of inhibiting ERα transactivation targeting its dimerization-a key process of ERα biological activity. Using in silico techniques, we first elucidated the hydrogen bond interactions involved in dimerization and identified three interfacial sequence motifs, where sequence I (DKITD) and sequence II (QQQHQRLAQ) of one monomer form hydrogen bonding with sequence II and sequence I of the second monomer, respectively, and sequence III (LSHIRHMSNK) hydrogen bonds with the same from the second monomer. Studying the structural stability and the binding affinity of the peptides derived from these sequence motifs, we found that an extended and ARG mutated version (LQQQHQQLAQ) of sequence II can act as a suitable template for designing peptidic inhibitors. It provides additional structural stability and interacts more strongly with ERα dimer interface groove formed by helices 9 and 10/11 and prevent ERα dimerization. Our result provides a novel therapeutic designing pipeline for ligand-independent inhibition of ERα.