Inhibitor peptide SNP‐1 binds to a soluble form of BST‐1/CD157 at a 2 : 2 stoichiometry

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Abstract
Recently we have identified a 15‐mer peptide, SNP‐1, by a random phage library that can bind to bone marrow stromal cell antigen‐1 (BST‐1)/CD157 [Sato, A., Yamamoto, S., Ishihara, K., Hirano, T. & Jingami, H. (1999) Biochem. J. 337, 491–496]. SNP‐1 inhibits BST‐1 ADP‐ribosyl cyclase activity uncompetitively with a Ki value of 180 ± 40 nm. In this study we analysed biophysically the SNP‐1 binding to a soluble form of BST‐1 (sBST‐1). Equilibrium binding data of wild‐type SNP‐1 from surface plasmon resonance studies gave a Kd value of 500 ± 35 nm. Titration calorimetry analysis showed that the binding reaction is exothermic at 20 °C. The values of Kd = 211 nm, enthalpy change, ΔH = –18.68 kcal·mol−1, and saturated molar ratio of bound SNP‐1 per sBST‐1, N = 0.8 mol·mol−1 were obtained. On the basis of the molecular masses of SNP‐1 and sBST‐1 calculated by analytical ultracentrifugation, the stoichiometry of the binding was determined to be 2 : 2. Electron microscopy also revealed the dimer form of sBST‐1. To delineate the core residue of SNP‐1 responsible for binding, each amino acid residue has been replaced by alanine. A region from amino acid residues 7–12 appeared to be critical for the SNP‐1 binding to sBST‐1. The substitution of the first residue, His, to Ala led to a reduction in binding, suggesting that the N‐terminal residue is also crucial.