CALCIUM-DEPENDENT BINDING OF S100C TO THE N-TERMINAL DOMAIN OF ANNEXIN-I

The annexin family of proteins is characterized by a conserved core do main that binds to phospholipids in a Ca2+-dependent manner. Each anne xin also has a structurally distinct N-terminal domain that may impart functional specificity. To search for cellular proteins that interact with the N-terminal domain of annexin I, we constructed a fusion prot ein consisting of glutathione S-transferase fused to amino acids 2-47 of human annexin I(GST AINT; AINT = annexin I N-terminal), Extracts fr om metabolically labeled A431 cells contained a single protein (M(r) s imilar to 10,000) that bound to GST-AINT in a Ca2+-dependent manner. A synthetic peptide corresponding to amino acids 2-18 of annexin I inhi bited the binding of the 10-kDa protein to GST-AINT with half-maximal inhibition occurring at similar to 15 mu M peptide. In cellular extrac ts, endogenous annexin I and the 10-kDa protein associated in a revers ible Ca2+ dependent manner. Experiments with other annexins and with N -terminal truncated forms of annexin I indicated that the 10-kDa prote in bound specifically to a site within the first 12 amino acids of ann exin I, The 10-kDa protein was purified from human placenta by hydroph obic and affinity chromatography. Amino acid sequence analysis indicat ed that the 10-kDa protein is the human homologue of S100C, a recently identified member of the S100 subfamily of EF-hand Ca2+-binding prote ins.