Several cytosolic proteins bind to secretory granule membranes in a Ca2+-dependent manner and thus may be involved in the mediation of membrane interactions during exocytosis. One of these proteins, calpactin, is a tetramer consisting of two heavy chains of relative molecular mass (Mr) 36K (p36) and two light chains of 10K (p10). We report here that calpactin promotes the Ca2+-dependent aggregation and fatty acid-dependent fusion of chromaffin granule membranes at a level of Ca2+ that is lower than that reported for other granule-aggregating proteins, and which parallels the Ca2+ requirement for secretion from permeabilized chromaffin cells. We found subunits of calpactin to be inactive in promoting granule aggregation. Two distinct 33K proteolytic fragments of p36, differing at their N termini, also promote granule aggregation but with different Ca2+ sensitivities from calpactin. These differences suggest that the N-terminal portion of p36 modulates the Ca2+/lipid binding sites in the core portion of p36 (ref.5).