CREB-binding protein (CBP) is a large, multi-domain protein that provides a multitude of binding sites for transcriptional coactivators. The site of interaction of the tumor suppressor p53 and the oncoprotein E1A with CBP/p300 has been identified with the third cysteine-histidine-rich (CH3) domain, which incorporates two zinc-binding motifs, ZZ and TAZ2. We show that these two domains fold independently and do not interact in solution. Our experiments demonstrate conclusively that the interaction of p53 and E1A with the CH3 domain resides exclusively in the TAZ2 domain, with no contribution from the ZZ domain. We report also the three-dimensional solution structure of the ZZ domain of murine CBP. The 52 residue ZZ domain contains two twisted antiparallel beta-sheets and a short alpha-helix, and binds two zinc ions. The identity of the zinc coordinating ligands was resolved unambiguously using NMR spectroscopy of the ZZ domain substituted with (113)Cd. One zinc ion is coordinated tetrahedrally via two CXXC motifs to four cysteine side-chains, and the second zinc ion is coordinated tetrahedrally by a third CXXC motif, together with an unusual HXH motif coordinating via the N(epsilon2) atom of His40 and the N(delta1) atom of His-42. The first zinc cluster of the ZZ domain is strictly conserved, whereas the second zinc cluster shows variability in the position of the two histidine residues, reflecting the wide variety of molecules that incorporate ZZ domains. The structure of the ZZ domain shows that it belongs to the family of cross-brace zinc finger motifs that include the PHD, RING, and FYVE domains; however, its biological function is unclear. Mapping of the positions of conserved residues onto the calculated structures reveals a face containing exposed aromatic and hydrophobic side-chains, while the opposite face contains a series of conserved charged or hydrophilic groups. These homologies suggest that the ZZ domain is involved in ligand binding or molecular scaffolding, with specificity provided by the variability of the sequence that contains the helix in the murine CPB ZZ domain structure.