Colosi P, Ogren L, Southard J N, Thordarson G, Linzer D I, Talamantes F
Department of Biochemistry, Molecular Biology, and Cell Biology, Northwestern University, Evanston, Illinois 60208.
Endocrinology. 1988 Dec;123(6):2662-7. doi: 10.1210/endo-123-6-2662.
Mouse placental lactogen-I (mPL-I) cDNA was inserted into a eukaryotic expression vector and introduced into Chinese hamster ovary cells. Cell lines that secrete high concentrations of mPL-I were isolated, and this glycoprotein was purified from the cell culture-conditioned medium. Recombinant mPL-I (mPL-Ir) is very similar to placental mPL-I (mPL-Ip) in its recognition by polyclonal antisera raised against either mPL-Ip or mPL-Ir, in displacing [125I]iodo-mPL-II from binding sites on mouse liver microsomal membranes, and in stimulating the synthesis of alpha-lactalbumin in primary cultures of mouse mammary epithelial cells. Structural comparison of mPL-Ir and mPL-Ip by sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicated that mPL-Ir comprises several proteins with mol wt ranging from 34.5-38K, while mPL-Ip consists of a similar set of proteins with mol wt ranging from 36.5-42K. Treatment of the two proteins with neuraminidase resulted in similar 2-4K decreases in mol wt. Treatment of mPL-Ip with peptide:N-glycosidase-F to remove asparagine-linked oligosaccharide chains resulted in the formation of 28K and 29K mol wt species, while treatment of mPL-Ir with the same enzyme yielded 28K and 28.5K mol wt products.
将小鼠胎盘催乳素-I（mPL-I）cDNA插入真核表达载体并导入中国仓鼠卵巢细胞。分离出分泌高浓度mPL-I的细胞系，并从细胞培养条件培养基中纯化该糖蛋白。重组mPL-I（mPL-Ir）在被针对mPL-Ip或mPL-Ir产生的多克隆抗血清识别、从小鼠肝微粒体膜上的结合位点置换[125I]碘-mPL-II以及刺激小鼠乳腺上皮细胞原代培养物中α-乳白蛋白的合成方面，与胎盘mPL-I（mPL-Ip）非常相似。通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳对mPL-Ir和mPL-Ip进行结构比较表明，mPL-Ir由几种分子量范围为34.5 - 38K的蛋白质组成，而mPL-Ip由一组类似的分子量范围为36.5 - 42K的蛋白质组成。用神经氨酸酶处理这两种蛋白质导致分子量类似地降低2 - 4K。用肽：N-糖苷酶-F处理mPL-Ip以去除天冬酰胺连接的寡糖链导致形成分子量为28K和29K的物种，而用相同酶处理mPL-Ir产生分子量为28K和28.5K的产物。
