Albani J R
Laboratoire de Biophysique Moléculaire, Université des Sciences et Technologies de Lille, B.P. 649, 59656 Villeneuve d'Ascq Cédex, France.
Biochim Biophys Acta. 1998 Oct 23;1425(2):405-10. doi: 10.1016/s0304-4165(98)00092-0.
Dynamics of the fluorescent Lens culinaris agglutinin-fluorescein complex (LCA-FITC) are studied in absence and in presence of two glycoproteins, lactotransferrin (LTF) and serotransferrin (STF). Glycans of the serotransferrin are not fucosylated, while those of the lactotransferrin have an alpha-1,6-fucose bound to the N-acetylglucosamine residue linked to the peptide chain, and an alpha-1,3-fucose bound to the N-acetyllactosamine residues. Interaction between the lectin and the two glycoproteins occurs via the carbohydrate residues. Affinity between LCA and LTF is 50 times higher than that between LCA and STF, as a result of the alpha-1, 6-fucose-LCA linkage. In the present work, we studied the effect of the tight bond between the alpha-1,6-fucose and LCA on the dynamics of the amino acids of the lectin, by fluorescence intensity quenching with iodide and by thermal intensity quenching. Fluorescence intensity quenching with iodide indicates that the bimolecular diffusion constant of iodide is 2.402+/-0.068x109 and 1. 160+/-0.090x109 M-1 s-1, when the interaction occurs with free fluorescein and with fluorescein bound to LCA, respectively. Binding of STF or LTF to the LCA-FITC complex yields a bimolecular diffusion constant of 1.675+/-0.06x109 and 1.155+/-0.087x109 M-1 s-1, respectively. Thermal intensity quenching does not occur for fluorescein free in solution while it is linear with temperature with a relative change of 0.656%, 0.889% and 0.488% for FITC-LCA, FITC-LCA-LTF and FITC-LCA-STF complexes, respectively. Fluorescence intensity quenching with iodide and thermal quenching experiments indicate that the dynamics of LCA increase as the result of the flexibility of the carbohydrate residues (case of STF-LCA complex), and the presence of the alpha-1,6-fucose inhibits the effect of the other carbohydrate residues as the result of the tight bond that exists between the fucose and the lectin (case of LTF-LCA complex).
在不存在和存在两种糖蛋白（乳铁传递蛋白（LTF）和血清转铁蛋白（STF））的情况下，研究了荧光菜豆凝集素 - 荧光素复合物（LCA - FITC）的动力学。血清转铁蛋白的聚糖不被岩藻糖基化，而乳铁传递蛋白的聚糖有一个与连接到肽链的N - 乙酰葡糖胺残基结合的α - 1,6 - 岩藻糖，以及一个与N - 乙酰乳糖胺残基结合的α - 1,3 - 岩藻糖。凝集素与这两种糖蛋白之间的相互作用通过碳水化合物残基发生。由于α - 1,6 - 岩藻糖 - LCA连接，LCA与LTF之间的亲和力比LCA与STF之间的亲和力高50倍。在本工作中，我们通过用碘化物进行荧光强度猝灭和热强度猝灭，研究了α - 1,6 - 岩藻糖与LCA之间的紧密结合对凝集素氨基酸动力学的影响。用碘化物进行荧光强度猝灭表明，当分别与游离荧光素和与LCA结合的荧光素相互作用时，碘化物的双分子扩散常数分别为2.402±0.068×10⁹和1.160±0.090×10⁹ M⁻¹ s⁻¹。STF或LTF与LCA - FITC复合物的结合分别产生双分子扩散常数为1.675±0.06×10⁹和1.155±0.087×10⁹ M⁻¹ s⁻¹。溶液中游离的荧光素不会发生热强度猝灭，而对于FITC - LCA、FITC - LCA - LTF和FITC - LCA - STF复合物，热强度猝灭与温度呈线性关系，相对变化分别为0.656%、0.889%和0.488%。用碘化物进行荧光强度猝灭和热猝灭实验表明，由于碳水化合物残基的柔韧性（STF - LCA复合物的情况），LCA的动力学增加，并且由于岩藻糖与凝集素之间存在紧密结合，α - 1,6 - 岩藻糖的存在抑制了其他碳水化合物残基的作用（LTF - LCA复合物的情况）。
