Calcium binding properties of gamma-crystallin - Calcium ion binds at the Greek key beta gamma-crystallin fold

The beta- and gamma -crystalline are closely related lens proteins that are members of the beta gamma -crystallin superfamily, which also include many non-lens members. Although beta -crystallin is known to be a calcium-bindi ng protein, this property has not been reported in gamma -crystallin. We ha ve studied the calcium binding properties of gamma -crystallin, and we show that it binds 4 mol eq of calcium with a dissociation constant of 90 muM. It also binds the calcium-mimic spectral probes, terbium and Stains-all. Ca lcium binding does not significantly influence protein secondary and tertia ry structures. We present evidence that the Greek key crystallin fold is th e site for calcium ion binding in gamma -crystallin. Peptides corresponding to Greek key motif of gamma -crystallin (42 residues) and their mutants we re synthesized and studied for calcium binding. These peptides adopt beta - sheet conformation and form aggregates producing beta -sandwich. Our result s with peptides show that, in Greek key motif, the amino acid adjacent to t he conserved aromatic corner in the "a" strand and three amino acids of the "d" strand participate in calcium binding. We suggest that the beta gamma superfamily represents a novel class of calcium-binding proteins with the G reek key beta gamma -crystallin fold as potential calcium-binding sites. Th ese results are of significance in understanding the mechanism of calcium h omeostasis in the lens.