Khan M I, Mathew M K, Balaram P, Surolia A
Biochem J. 1980 Nov 1;191(2):395-400. doi: 10.1042/bj1910395.
The binding of Ricinus communis (castor-bean) agglutinin 1 to saccharides was studied by equilibrium dialysis and fluorescence polarization by using the fluorescently labelled sugar 4-methylumbelliferyl beta-D-galactopyranoside. No appreciable change in ligand fluorescence of 4-methylumbelliferyl beta-D-galactopyranoside was considerably polarized on its binding to the lectin. The association constants obtained by Scatchard analysis of equilibrium-dialysis and fluorescence-polarization data do not differ much from each other, and at 25 degrees C, Ka = 2.4 (+/- 0.2) X 10(4)M-1. These values agree reasonably well with that reported in the literature for Ricinus agglutinin 1. The number of binding sites obtained by the different experimental procedures is 1.94 +/- 0.1 per molecule of 120 000 daltons and is equal to the reported value of 2. The consistency in the values of Ka and number of binding sites indicate the absence of additional subsites on Ricinus agglutinin 1 for its specific sugars. In addition, the excellent agreement between the binding parameters obtained by equilibrium dialysis and fluorescence polarization indicate the potential of ligand-fluorescence-polarization measurements in the investigation of lectin-sugar interactions.
通过平衡透析法并利用荧光标记的糖4-甲基伞形酮基β-D-吡喃半乳糖苷，采用荧光偏振技术研究了蓖麻凝集素1与糖类的结合情况。4-甲基伞形酮基β-D-吡喃半乳糖苷的配体荧光在与凝集素结合时没有明显变化，但偏振程度显著增加。通过对平衡透析和荧光偏振数据进行Scatchard分析得到的缔合常数彼此相差不大，在25℃时，Ka = 2.4（±0.2）×10⁴M⁻¹。这些值与文献中报道的蓖麻凝集素1的值相当吻合。通过不同实验方法得到的结合位点数为每120000道尔顿分子1.94±0.1个，与报道的2个值相等。Ka值和结合位点数的一致性表明蓖麻凝集素1对于其特定糖类不存在额外的亚位点。此外，平衡透析法和荧光偏振法得到的结合参数之间的良好一致性表明，配体荧光偏振测量在研究凝集素-糖相互作用方面具有潜力。
