An extended hydrophobic core induces EF-hand swapping

The structure of calbindin D-9k With two substitutions was determined by X-ray crystallography at 1.8-Angstrom resolution. Unlike wild-type calbindin D9k, which is a monomeric protein with two EF-hands, the structure of the mutated calbindin D-9k reveals an intertwined dimer. In the dimer, two EF-hands of the monomers have exchanged places, and thus a 3D domain-swapped dimer has been formed. EF-hand I of molecule A is packed toward EF-hand II of molecule B and vice versa. The formation of a hydrophobic cluster, in a region linking the EF-hands, promotes the conversion of monomers to 3D domain-swapped dimers. We propose a mechanism by which domain swapping takes place via the apo form of calbindin D-9k. Once formed, the calbindin D-9k dimers are remarkably stable, as with even larger misfolded aggregates like amyloids. Thus calbindin D-9k dimers cannot be converted to monomers by dilution. However, heating can be used for conversion, indicating high energy barriers separating monomers from dimers.