棘托竹荪凝集素的纯化及其生化特性

Purification and characterization of the lectin from Dictyophora echinovolvata

棘托竹荪(DictyophoraechinovolvataZang,ZhengetHu)子实体经生理盐水抽提、硫酸铵沉淀、DEAE Sepharose和SephadexG 100柱层析纯化得到棘托竹荪凝集素(DEL)。经PAGE显示单一条带,相对分子质量为38000,其亚基相对分子质量为18900;不含中性糖,IEF PAGE测得其等电点为4.21。该凝集素对供试的4种血型人血和6种动物血的红细胞具有凝集作用,也能凝集小鼠淋巴细胞和小鼠S180肉瘤细胞,对兔红细胞的凝集作用可被乳糖和果糖所抑制。DEL含有15种氨基酸,其中天冬氨酸、谷氨酸、甘氨酸和缬氨酸含量较高;N 末端为丙氨酸。DEL对热不稳定,经50℃处理10min,活性明显降低;在pH4.00～pH10.14范围内较稳定;其凝血活性依赖于Mn2+和Ca2+,Mg2+和Zn2+则无影响。DEL对小鼠腹腔注射的半致死量为1180mg·kg-1。

The lectin DEL from the fruiting bodies of Dictyophora echinovolvata Zang, Zheng et Hu was isolated and purified by physiological saline extraction, precipitation of 40% (NH_(4))_(2)SO_(4) and DEAE-Sepharose, Sephadex G-100 chromatography purification. It turned out to be a single band in PAGE identification. The relative molecular mass of DEL was 38 000 and the subunit 18 900, its pI was about 4.21. This lectin agglutinated with 4 types of human erythrocytes and 6 types of animal erythrocytes, it also agglutinated with the lymphocyte cells and S_(180) tumor cells from mouse. The hemagglutination of the lectin on rabbit erythrocytes was inhibited by lactose and fructose. It contained 15 kinds of amino acid, in which the contents of aspartic acid, glutamic acid, glycine and valine were relatively high. The N-terminal of DEL was alanine. DEL was thermal-unstable, its agglutination activity decreased obviously after being heated 10 min at 50℃, but had high activity within pH 4.00-pH 10.14. Its activity was depended on Mn^(2+) and Ca^(2+), not on Mg^(2+) and Zn^(2+). DEL was toxic to mouse by celiac injection with a LD_(50) of 1 180 mg·kg^(-1).