The 14-3-3 protein binds its target proteins with a common site located towards the C-terminus.

The 14-3-3 protein family binds a variety of proteins in cell-signaling pathways, but the structural elements necessary for the ligand binding are poorly understood. Here we demonstrate that the 'box-1' region, which spans residues 171-213 in the eta-isoform and was previously identified as the binding site of 14-3-3 to the phosphorylated tryptophan hydroxylase, plays a critical role in the interaction with many target proteins. Using a series of truncated 14-3-3 mutants, we show that the mutant 167-213 carrying box-1 binds bacurovirus-expressed Raf-1 and Bcr protein kinases to the similar extent as the full-length 14-3-3 in a phosphorylation-dependent manner, while the mutants lacking this region abolish the binding activity. Furthermore, the box-1 region also appears essential for binding of 14-3-3 to more than 40 phosphoproteins found in the brainstem extract. These results suggest that the box-1 region, consisting of helices 7 and 8 in the tertiary structure, is a common structural element whereby the 14-3-3 protein binds many, if not all, target proteins.