Faculty of Pharmaceutical Sciences, Josai University, Sakado, Saitama, Japan.
Biol Pharm Bull. 2011;34(6):929-32. doi: 10.1248/bpb.34.929.
LEC-1 is the first tandem repeat-type galectin isolated from an animal system; this galectin has two carbohydrate recognition domains in a single polypeptide chain. Because its two lectin domains have different sugar-binding profiles, these domains are thought to interact with different carbohydrate ligands. In our previous study, we showed that a mutant of LEC-1 in which a cysteine residue was introduced at a unique position in the N-terminal lectin domain (Nh) can be cross-linked with a model glycoprotein ligand, bovine asialofetuin, by using a bifunctional photoactivatable cross-linking reagent, benzophenone-4-maleimide. In the present work, we applied the same procedure to the C-terminal lectin domain (Ch) of LEC-1. Cross-linked products were formed in the cases of two mutants in which a cysteine residue was introduced at Lys¹⁷⁷ and Ser²⁶⁸, respectively. This method is very useful for capturing and assigning endogenous ligand glycoconjugates with relatively low affinities to each carbohydrate recognition domain of the whole tandem repeat-type galectin molecule.
LEC-1 是从动物系统中分离出的第一个串联重复型半乳糖凝集素；这种半乳糖凝集素在单个多肽链中有两个碳水化合物识别结构域。由于其两个凝集素结构域具有不同的糖结合谱，因此这些结构域被认为与不同的碳水化合物配体相互作用。在我们之前的研究中，我们表明，在 LEC-1 的 N 端凝集素结构域（Nh）的独特位置引入半胱氨酸残基的突变体可以与模型糖蛋白配体牛去唾液酸胎球蛋白通过使用双功能光活化交联试剂苯并酮-4-马来酰亚胺交联。在本工作中，我们将相同的程序应用于 LEC-1 的 C 端凝集素结构域（Ch）。在分别将半胱氨酸残基引入 Lys¹⁷⁷ 和 Ser²⁶⁸ 的两种突变体中形成了交联产物。该方法非常有用，可以捕获和分配与整个串联重复型半乳糖凝集素分子的每个碳水化合物识别结构域具有相对较低亲和力的内源性配体糖缀合物。
