Katyal S L, Singh G
Biochim Biophys Acta. 1984 Jul 26;794(3):411-8. doi: 10.1016/0005-2760(84)90007-9.
Apoproteins of Mr 38 000, 32 000 and 26 000 are found in surfactant isolated from rat lungs. The surfactant isolated from monkey lungs, on the other hand, contains the 38 kDa apoprotein and not the 32 and 26 kDa apoproteins. These preparations of pulmonary surfactant contain, in addition, several serum proteins. We have used a combination of salt- and sucrose-density gradient centrifugations to isolate and further purify surfactant from the washings of rat lungs. Thus, a preparation of pulmonary surfactant was obtained which contained exclusively the 38, 32, 26 and 10-12 kDa apoproteins, and which was rich in phosphatidylcholine and phosphatidylglycerol. Using an immunoassay and an immunoblotting technique, it was established that the 38, 32 and 26 kDa apoproteins are not serum proteins. The surfactant apoproteins of rat and monkey were further subjected to the high-resolution of isoelectric focusing. Thus, rat surfactant apoproteins resolved into 11 bands in the pH range 4.64-5.53. A second-dimensional electrophoresis in a sodium dodecyl sulfate system led to the migration of the 11 bands, separated by first-dimensional isoelectric focusing, into three distinct groups with apparent molecular weights of 38 000, 32 000 and 26 000, respectively. Upon isoelectric focusing, the apoproteins from monkey lung surfactant also separated into several bands in the pH range 5.18-5.82. After electrophoresis in the second dimension as above, these bands migrated as a single group with an apparent molecular weight of 38 000. Neuraminidase treatment of rat surfactant apoproteins, and subsequent IEF, led to the disappearance of several low-pI variants with a concomitant increase in the amounts of higher-pI variants. Thus, the sialic acid content of surfactant apoproteins accounts for, in large part, the observed charge heterogeneity.
在从大鼠肺中分离出的表面活性剂中发现了分子量为38000、32000和26000的载脂蛋白。另一方面，从猴肺中分离出的表面活性剂含有38kDa的载脂蛋白，而不含32kDa和26kDa的载脂蛋白。这些肺表面活性剂制剂还含有几种血清蛋白。我们使用盐和蔗糖密度梯度离心相结合的方法，从大鼠肺洗液中分离并进一步纯化表面活性剂。因此，获得了一种肺表面活性剂制剂，其仅含有38、32、26和10 - 12kDa的载脂蛋白，并且富含磷脂酰胆碱和磷脂酰甘油。使用免疫测定和免疫印迹技术确定，38、32和26kDa的载脂蛋白不是血清蛋白。大鼠和猴的表面活性剂载脂蛋白进一步进行了等电聚焦的高分辨率分析。因此，大鼠表面活性剂载脂蛋白在pH范围4.64 - 5.53内分离成11条带。在十二烷基硫酸钠系统中进行的二维电泳导致通过一维等电聚焦分离的11条带迁移成三个不同的组，其表观分子量分别为38000、32000和26000。等电聚焦时，猴肺表面活性剂的载脂蛋白在pH范围5.18 - 5.82内也分离成几条带。在上述二维电泳后，这些条带作为一个单一的组迁移，表观分子量为38000。用神经氨酸酶处理大鼠表面活性剂载脂蛋白，随后进行IEF，导致几个低pI变体消失，同时高pI变体的量增加。因此，表面活性剂载脂蛋白的唾液酸含量在很大程度上解释了观察到的电荷异质性。
