Kobayashi R, Reeve J R, Walsh J H
Biochem J. 1985 Aug 1;229(3):611-9. doi: 10.1042/bj2290611.
Canine epidermal growth factor (EGF)/urogastrone was partially purified from dog urine by fractional precipitation with (NH4)2SO4, ion-exchange chromatography with DEAE-cellulose DE-52, gel filtration with Sephadex G-50, and a second DE-52 chromatography, to yield receptor-competing activity equivalent to 13 micrograms of standard mouse EGF/litre of starting urine. The purification was monitored by a competitive radioreceptor assay using fixed monolayers of A431 cells. The partially purified canine EGF/urogastrone demonstrated a growth-stimulating activity in 3T3 mouse fibroblast cells as potent as mouse EGF. Analysis by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis revealed one major peptide component with an Mr similar to that of mouse EGF, and two minor peptides of slightly higher Mr. The major peptide component was isolated after reduction and its amino acid composition was determined.
通过用硫酸铵分级沉淀、DEAE - 纤维素DE - 52离子交换色谱、Sephadex G - 50凝胶过滤以及第二次DE - 52色谱，从犬尿中部分纯化犬表皮生长因子（EGF）/尿抑胃素，得到的受体竞争活性相当于每升起始尿液中13微克标准小鼠EGF。使用A431细胞固定单层的竞争性放射受体测定法监测纯化过程。部分纯化的犬EGF/尿抑胃素在3T3小鼠成纤维细胞中表现出与小鼠EGF一样强的生长刺激活性。十二烷基硫酸钠/聚丙烯酰胺凝胶电泳分析显示有一个主要肽成分，其相对分子质量（Mr）与小鼠EGF相似，还有两个Mr略高的次要肽。主要肽成分经还原后分离出来并测定了其氨基酸组成。
