Nagamatsu Y, Oka T
Biochem J. 1980 Jan 1;185(1):227-37. doi: 10.1042/bj1850227.
alpha-Lactalbumin was purified to apparent homogeneity from mouse milk by combined use of gel filtration, chromatography on DEAE-cellulose and hydroxyapatite, and concanavalin A-Sepharose affinity chromatography. Mouse alpha-lactalbumin exists in several species with different charges and in two molecular-size forms. The smaller form, which constituted over 90% of total alpha-lactalbumin, included two major and two minor species, each of which showed different electrophoretic mobility on polyacrylamide-gel electrophoresis, but gave the same single band on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis in two different buffer systems and over the range 10-15% acrylamide concentrations. The molecular weight was estimated as 14 100. The two major species of the smaller form had the same amino acid composition and contained no significant amount of carbohydrate. The larger form of alpha-lactalbumin, consisting of two species with different charges, was present in a small amount (less than 10%) in the milk and was isolated by its ability to interact with concanavalin A-Sepharose. Each of the two species also gave the same single band of apparent mol.w.t 18 500 on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. However, this value may be anomalous, since this larger form appears to be glycosylated, and glycoproteins can behave anomalously on sodium dodecyl sulphate/polyacrylamide gels by binding less sodium dodecyl sulphate. All species of mouse alpha-lactalbumin from milk were active in the lactose synthase reaction and showed identical immunological properties, as determined by the mono-specific antibody prepared against the small major species. The presence of both the larger and the smaller forms, each in a percentage concentration similar to that found in milk, was also demonstrated in alpha-lactalbumin induced by hormones in organ cultureof pregnant-mouse mammary gland.
通过联合使用凝胶过滤、DEAE - 纤维素和羟基磷灰石层析以及伴刀豆球蛋白A - 琼脂糖亲和层析，从小鼠乳汁中纯化出了外观上均一的α - 乳白蛋白。小鼠α - 乳白蛋白以几种带不同电荷的形式以及两种分子大小的形式存在。较小的形式占总α - 乳白蛋白的90%以上，包括两个主要种类和两个次要种类，每一种在聚丙烯酰胺凝胶电泳上都显示出不同的电泳迁移率，但在两种不同缓冲体系以及10 - 15%丙烯酰胺浓度范围内的十二烷基硫酸钠/聚丙烯酰胺凝胶电泳上给出相同的单一条带。分子量估计为14100。较小形式的两个主要种类具有相同的氨基酸组成，且不含大量碳水化合物。α - 乳白蛋白的较大形式由两种带不同电荷的种类组成，在乳汁中含量较少（不到10%），通过其与伴刀豆球蛋白A - 琼脂糖相互作用的能力被分离出来。这两种种类在十二烷基硫酸钠/聚丙烯酰胺凝胶电泳上也都给出了表观分子量为18500的相同单一条带。然而，这个值可能是异常的，因为这种较大形式似乎是糖基化的，并且糖蛋白在十二烷基硫酸钠/聚丙烯酰胺凝胶上可能会因结合较少的十二烷基硫酸钠而表现异常。乳汁中的所有小鼠α - 乳白蛋白种类在乳糖合酶反应中都具有活性，并且如针对小的主要种类制备的单特异性抗体所测定的那样，显示出相同的免疫特性。在怀孕小鼠乳腺器官培养中由激素诱导产生的α - 乳白蛋白中也证实了较大和较小形式的存在，且每种形式的百分比浓度与乳汁中的相似。
