The cellular prion protein (PrP) binds to the C-terminus of Bcl-2 but not Bax. Therefore, we examined whether the C-terminus of Bcl-2 was important for other homomeric and heteromeric protein-protein interactions. Using the yeast two hybrid system and co-immunoprecipitation, three sites of homomeric interactions were identified within Bcl-2. The carboxy terminal 37 amino acids selectively homodimerized. Two additional regions of Bcl-2 (residues 1-129 and 126-200) interacted with each other, but not themselves permitting both intra- and intermolecular association. In addition, we analyzed heteromeric interactions of Bcl-2 with PrP and two Bcl-2 related proteins, Bax and A1. The domain requirements for binding of those three proteins to Bcl-2 were different from one another. Bax binding required almost the entire Bcl-2 molecule, while A1 bound to the amino terminal region (residues 1-82). PrP associated with the carboxy terminus of Bcl-2 (amino acids 200-236). These data suggest configurational models for Bcl-2 containing complexes. First, Bcl-2 may exist as both heterodimers and heteromultimers. Second, molecules such as Bax and A1 may serve to cap chains of Bcl-2 homodimers by interacting with dimerization domains in the extramembrane region. PrP may disrupt chains of Bcl-2 molecules at the homomeric association site in the transmembrane region.