The carbohydrate-binding domain of Bauhinia purpurea lectin (BPA), a galactose and lactose-binding lectin, was investigated from Asp-N endoproteinase digests of BPA. A peptide which interacted with lactose was purified by means of lactose-Sepharose column chromatography. It consisted of 9 amino acids and its amino acid sequence was DTWPNTEWS. The chemical synthesis of this nonapeptide was carried out by the solid-phase method and the synthetic peptide was found to exhibit lac-tose-specific binding activity in the presence of calcium. Similar studies were carried out on several anti-H (O) lectins and the peptides which showed affinity were also found as a part of the metal-binding region of these lectins. Subsequently, we constructed a chimeric lectin gene in which nonapeptide sequence was replaced by the corresponding region of the mannose-binding Lens culinaris lectin. The chimeric lectin expressed in E. coli was found to bind α-mannosyl-bovine serum albumin. These results are the first to describe the presence of a “variable-binding region” in legume lectins that detemine their carbohydrate-binding specificity