天花粉同工凝集素－1双链的拆分和鉴定

天花粉同工凝集素－１经巯基乙醇还原，碘代乙酰胺保护，其链间二硫键被打开，但仍非共价结合在一起。我们利用尿素变性的Ｑ－Ｓｅｐｈａｒｏｓｅ离子交换层忻分离了此凝集素的两条链。氨基酸组成测定与其他３种肽链作一比较，它们都含有较多的酸性和羟基氨基酸。蛋白质印迹显示ＴＫＬ的抗血清不仅能与ＴＫＬ－１的两条链分别反应，也能与天花粉毒蛋白及蓖麻毒蛋白的Ａ链起作用。溴化氰裂解的ＳＤＳ－ＰＡＧＥＲ肽谱表明天花粉凝集素的两条链与天花粉毒蛋白含有类似的裂解片段，在分子量１６ｋｄ左右有相同的电泳条带。ＴＫＬ－１两亚基的Ｎ末端序列已经测定，同源性比较发现其３３ｋｄ亚基的Ｎ末端序列与天花粉毒蛋白、蓖麻毒蛋白的一些肽段类似。迄今已有的证据表明ＴＫＬ与ＴＣＳ等是一些非常相关的蛋白质。

Separation and Characterization of the Two Chains of Trichosanthes kirilowii Isolectins 1

The two chains of TKL-1 were bound tightly after reduction and alkylation. They could be separated from each other in 8M urea using fast flow Q-Sepharose chromatography. Their amino acid compositions were analysed and compared with three other peptides: trichosanthin, ricin A chain and ricin B chain. They were all rich in acidic and hydroxyl amino acids. Western blot anslysis of the leduced TKL-1, ricin and TCS showed that antibodies against TKL could react with both the two chains of TKL-1, TCS and ricin A chain. SDS-PAGE peptide mapping of the two chains of TKL-1 and TCS after CNBr cleavage resulted in one similar band of molecular weight about 16kd. N-Terminal sequences of the two chains of TKL-1 were determined and exhibited some homologies with TCS, ricin A, B chains. Therefore all the evidence we have now suggests that these three proteins are structurally related.