The C-terminal cytoplasmic tail of polycystin-2 (PC2/TRPP2), a Ca 2+ -permeable channel, is frequently mutated or truncated in autosomal dominant polycystic kidney disease. We have previously shown that this tail consists of three functional regions: an EF-hand domain (PC2-EF, 720–797), a flexible linker (798–827), and an oligomeric coiled coil domain (828–895). We found that PC2-EF binds Ca 2+ at a single site and undergoes Ca 2+ -dependent conformational changes, suggesting it is an essential element of Ca 2+ -sensitive regulation of PC2 activity. Here we describe the NMR structure and dynamics of Ca 2+ -bound PC2-EF. Human PC2-EF contains a divergent non-Ca 2+ -binding helix-loop-helix (HLH) motif packed against a canonical Ca 2+ -binding EF-hand motif. This HLH motif may have evolved from a canonical EF-hand found in invertebrate PC2 homologs. Temperature-dependent steady-state NOE experiments and NMR R 1 and R 2 relaxation rates correlate with increased molecular motion in the EF-hand, possibly due to exchange between apo and Ca 2+ -bound states, consistent with a role for PC2-EF as a Ca 2+ -sensitive regulator. Structure-based sequence conservation analysis reveals a conserved hydrophobic surface in the same region, which may mediate Ca 2+ -dependent protein interactions. We propose that Ca 2+ -sensing by PC2-EF is responsible for the cooperative nature of PC2 channel activation and inhibition. Based on our results, we present a mechanism of regulation of the Ca 2+ dependence of PC2 channel activity by PC2-EF.