泥蚶(Tegillarca granosa Linnaeus)血淋巴液凝集素的分离纯化及其性质研究

泥蚶是一种重要的海产经济贝类,其血淋巴液经硫酸铵二步分级沉淀后,再经Sephadex G- 100凝胶过滤和Sepharose 4B亲和层析纯化制得泥蚶血淋巴液凝集素。经测定,该凝集素分子量约为123Kda,为两个亚基的蛋白质,其相对分子量分别为15 KDa和16 KDa,分子中含5．02％的糖。在氨基酸组成中,天门冬氨酸(Asp)含量最高,其次是谷氨酸(Glu)和组氨酸(His),不含蛋氨酸(Met)。泥蚶血淋巴液凝集素对多种天然或经酶修饰的人或动物红细胞具有不同的凝集作用,其中对兔红细胞的凝集活性最大。半乳糖和乳糖对其凝集活性具有抑制作用。凝集活性依赖于Ca2 ,在pH7．0较稳定,热稳定性不高,在30℃-70℃时凝集效价由原来的25下降为21,当温度超过80℃以后,凝血活性完全丧失。

PURIFICATION AND CHARACTERIZATION OF HEMOLYMPH LECTIN FROM TEGILLARCA GRANOSA LINNAEUS

Agglutinin with hemagglutinating activity was found in hemolymph of Tegillarca granosa Linnaeus. By extraction, fraction with saturated ammonium sulfate precipitation, gel filtration on Sephadex G-100 and followed by affinity chromatography on Sepharose 4B, an agglutinin was purified from the hemolymph of T. granosa. The purified lectin contains 5.02% neutral saccharide. The molecular weights is about 123 KDa, which showed 2 bands on SDS-PAGE, and the relative molecular weights of these subunits were 15 KDa and 16 KDa, respectively. In its amino acid composition, Asp has the highest content, followed by Glu and His, but there is a lack of Met. Assays for erythrocyte agglutination and sugar inhibition were done in microtiter plates. The result indicared that TGL-hemolymph could agglutinate erythrocytes from the natural and enzyme-treated erythrocytes of human and animals, and the highest activity was found in the agglutination with rabbit erythrocytes. The agglutinic activity of TGL-hemolymph was inhibited by lactose and D-galactose. The hemagglutination activity of TGL-hemolymph depended on Ca2+, and it showed the highest hemagglutination activity in pH 7.0. Thermostability was not high. The agglutinating activities changed from 2(5) to 2(1) between 30-70 degrees C, and was completely destroyed beyond 80 degrees C.