Ooi L S, Ng T B, Sun S S, Ooi V E
Department of Biochemistry, Chinese University of Hong Kong, Shatin, New Territories.
J Protein Chem. 2000 Feb;19(2):163-8. doi: 10.1023/a:1007042902391.
Three mannose-specific lectins exhibiting considerable similarities in NH2-terminal amino acid sequence were isolated from leaves of the Chinese daffodil Narcissus tazetta (Family Amaryllidaceae). The purification protocol involved extraction with an aqueous buffer, anion exchange chromatography on DEAE-cellulose using stepwise elution with increasing salt concentrations, affinity chromatography on mannose-agarose, and FPLC-gel filtration on Superose 12. From the peak unadsorbed on DEAE-cellulose, and two peaks adsorbed on the ion exchanger and eluted respectively with 0.2 M Tris-HCl buffer and 0.5 M NaCl, were prepared fractions which yielded isolectins 1, 2, and 3 after adsorption on mannose-agarose and FPLC-gel filtration. All three isolectins were homodimers with a molecular weight of 26 kDa. The lectin unadsorbed on DEAE-cellulose had the lowest, while the most strongly adsorbed lectin had the highest hemagglutinating activity.
从中国水仙(石蒜科)叶片中分离出三种在氨基末端氨基酸序列上具有相当相似性的甘露糖特异性凝集素。纯化方案包括用水性缓冲液提取、在DEAE - 纤维素上进行阴离子交换色谱(使用逐步增加盐浓度的洗脱方式)、在甘露糖 - 琼脂糖上进行亲和色谱以及在Superose 12上进行快速蛋白质液相色谱 - 凝胶过滤。从DEAE - 纤维素上未吸附的峰以及离子交换剂上吸附的两个峰(分别用0.2 M Tris - HCl缓冲液和0.5 M NaCl洗脱)中制备了馏分,这些馏分在吸附到甘露糖 - 琼脂糖上并经过快速蛋白质液相色谱 - 凝胶过滤后产生了凝集素1、2和3。所有三种同工凝集素均为同型二聚体,分子量为26 kDa。未吸附在DEAE - 纤维素上的凝集素血凝活性最低,而吸附最强的凝集素血凝活性最高。