Heavy-atom derivatives in lipidic cubic phases: results on hen egg-white lysozyme tetragonal derivative crystals with Gd-HPDO3A complex.

Gd-HPDO3A, a neutral gadolinium complex, is a good candidate for obtaining heavy-atom-derivative crystals by the lipidic cubic phase crystallization method known to be effective for membrane proteins. Gadolinium-derivative crystals of hen egg-white lysozyme were obtained by co-crystallizing the protein with 100 mM Gd-HPDO3A in a monoolein cubic phase. Diffraction data were collected to a resolution of 1.7 A using Cu Kalpha radiation from a rotating-anode generator. Two binding sites of the gadolinium complex were located from the strong gadolinium anomalous signal. The Gd-atom positions and their refined occupancies were found to be identical to those found in derivative crystals of hen egg-white lysozyme obtained by co-crystallizing the protein with 100 mM Gd-HPDO3A using the hanging-drop technique. Moreover, the refined structures are isomorphous. The lipidic cubic phase is not disturbed by the high concentration of Gd-HPDO3A. This experiment demonstrates that a gadolinium complex, Gd-HPDO3A, can be used to obtain derivative crystals by the lipidic cubic phase crystallization method. Further studies with membrane proteins that are known to crystallize in lipidic cubic phases will be undertaken with Gd-HPDO3A and other Gd complexes to test whether derivative crystals with high Gd-site occupancies can be obtained.