Mitra D, Sarkar M, Allen A K
Biochem J. 1987 Mar 1;242(2):331-8. doi: 10.1042/bj2420331.
The cold agglutinin from the albumin gland of the snail Achatina fulica was purified to homogeneity by using sheep gastric mucin-Sepharose 4B as affinity column followed by gel filtration on Bio-Gel P-300. The homogeneity was checked by alkaline gel electrophoresis, immunodiffusion and immunoelectrophoresis. The purified cold agglutinin is a glycoprotein of native M2 220,000 consisting of three non-covalently bound subunits of Mr 84,000, 74,000 and 62,000 and having a pI value of 4.5. The predominant amino acids are aspartic acid and glutamic acid (or amides) and serine, which account for 39% of the residues. About 3% of the residues are half-cystine. The lectin is a glycoprotein with about 30.7% carbohydrate, the most abundant sugars being galactose, N-acetylgalactosamine and N-acetylglucosamine. Mannose, xylose and fucose are also present. The inhibition of agglutination of human umbilical-cord erythrocytes by the cold agglutinin is specific for methyl beta-D-galactoside and also for glycolipids present on cord erythrocytes. The c.d. data show only negative ellipticity values in the far-u.v. region for the protein at various concentrations and temperatures and also in the presence of the hapten lactose (at different concentrations), indicating the presence of a random-coil conformation in the agglutinin that varies according to temperature.
以羊胃粘蛋白 - 琼脂糖凝胶4B为亲和柱，随后在Bio - Gel P - 300上进行凝胶过滤，将褐云玛瑙螺白蛋白腺中的冷凝集素纯化至均一。通过碱性凝胶电泳、免疫扩散和免疫电泳检查其均一性。纯化的冷凝集素是一种天然M2 220,000的糖蛋白，由分子量为84,000、74,000和62,000的三个非共价结合亚基组成，pI值为4.5。主要氨基酸是天冬氨酸、谷氨酸（或酰胺）和丝氨酸，占残基的39%。约3%的残基是半胱氨酸。该凝集素是一种糖蛋白，含约30.7%的碳水化合物，最丰富的糖是半乳糖、N - 乙酰半乳糖胺和N - 乙酰葡糖胺。也存在甘露糖、木糖和岩藻糖。冷凝集素对人脐带红细胞凝集的抑制作用对β - D - 甲基半乳糖苷以及脐带红细胞上存在的糖脂具有特异性。圆二色性数据显示，在不同浓度和温度下以及存在半抗原乳糖（不同浓度）时，该蛋白在远紫外区域仅呈现负椭圆率值，表明凝集素中存在随温度变化的无规卷曲构象。
