X-ray absorption fine structure investigation of the zinc transition metal binding site of Zn concanavalin A in solution and in the crystal.

We report details on measurements by the X-ray absorption fine structure (XAFS) technique of the conformational changes around the transition metal binding site (S1) of the protein concanavalin A induced by crystallization when that site is occupied by Zn. A change from hexa- to tetracoordination occurs at the S1 site on crystallization when the calcium-binding site (S2) is occupied by a calcium atom. When the S2 site is unoccupied, the Zn is pentacoordinated both in solution and in the crystal. The average distance to the coordination shell increases with coordination number as expected. Conformational changes are detected up to 4.5 A from the Zn, the limit of sensitivity of the XAFS technique. When the Zn is hexacoordinated, the ligands around the Zn, as determined by XAFS, are consistent with the crystal structure determination results of five oxygens and one nitrogen. The atom that is released in the tetracoordinated Zn. decreases to five is an oxygen atom, and, in addition, the nitrogen is released in the tetracoordinated Zn. Thus, when S2 is emptied, the protein gains a ligand about the Zn site in the crystal and loses one in solution. These results provide direct evidence that the protein conformation can be altered by the intermolecular forces of crystallization.