PHOSPHORYLATION AND CALCIUM-BINDING PROPERTIES OF AN ARABIDOPSIS GF14BRAIN PROTEIN HOMOLOG

Arabidopsis GF14 omega was originally described because of its apparen t association with a DNA-protein complex; it is a member of the 14-3-3 kinase regulatory protein family that is conserved throughout eukaryo tes. Here, we demonstrated that recombinant GF14 omega is expressed in Escherichia coli as a dimer. Blot binding and electrophoretic mobilit y shift analyses indicated that GF14 omega binds calcium. Equilibrium dialysis further demonstrated that GF14 omega binds an equimolar amoun t of calcium with an apparent binding constant of 5.5 x 10(4) M(-1) un der physiological conditions. The C-terminal domain, which contains a potential EF hand motif, is responsible for the calcium binding. The C -terminal domain also cross-reacted with the anti-GF14 omega monoclona l antibody. In addition, GF14 omega is phosphorylated by Arabidopsis p rotein kinase activity at a serine residue(s) in vitro. Therefore, GF1 4 omega protein has biochemical properties consistent with potential s ignaling roles in plants. The presence of a potential EF hand-like mot if in the highly conserved C terminus of 14-3-3 proteins together with the calcium-dependent multiple functions attributed to the 14-3-3 pro teins indicate that the C terminus EF hand is a common functional elem ent of this family of proteins.