Estrogen dependent breast cancers require the transcriptional activation activity of the estrogen receptor alpha (ERα). These account for approximately 70% of all breast cancers. In response to estrogen binding, a set of genes is activated by ERα. This facilitates growth and propagation of the cancer cells. The important calcium-binding signaling protein calmodulin (CaM) binds ERα, and, recently, it was demonstrated that CaM is essential for activation of ERα transcriptional activity. Here we present progress towards defining the CaM binding region of ERα and the structural changes in CaM upon interaction with ERα, and towards understanding potential cellular mechanisms for mediating activation of ERα by CaM. As a first step towards elucidating the mechanism of ERα activation by CaM, we have produced a segment of the ERα protein (residues 241-320) as a thioredoxin fusion and demonstrated binding to CaM. A short, 25 amino acid section of this segment of ERα, suspected to comprise the CaM binding sequence, was produced, as were three derivatives containing modifications known to occur naturally. Fluorescence titration experiments demonstrated that these peptides all bind to CaM, and they all bind with rare 2:1 (peptide:CaM) stoichiometries. Their relative binding affinities are consistent with established principles for CaM interactions with target domains. The complex of CaM with the wild-type peptide was studied using NMR spectroscopy. The chemical shifts of CaM were assigned using triple resonance methods. The changes in chemical shifts upon peptide binding suggest an altered binding mode relative to typical complexes of CaM with target peptides.