Yousef I M, Murray R K
Can J Biochem. 1978 Jul;56(7):713-21. doi: 10.1139/o78-107.
Plasma membrane and bile canalicular membrane fractions were prepared from rat liver using NaHCO3, NaHCO3--CaCl2, and K2HPO4-KH2PO4 buffers (all at pH 7.4). The amount (expressed as milligrams protein per gram liver) of plasma membrane fraction exceeded the amount of bile canalicular membrane fraction using each of these three media; the use of NaHCO3-CaCl2 afforded a substantially higher yield of both types of membranes. The two membrane fractions exhibited complex patterns of polypeptides (greater than 30) on sodium dodecyl sulphate (SDS) polyacrylamide gel electrophoresis. Several reproducible differences in polypeptide patterns were observable between the two membrane fractions; in particular, components possibly corresponding to the heavy chain of myosin and to action were prominent in the bile canalicular membrane fraction. The effects of incubation in the above three buffers and in Tris--HCl (pH 7.4) on the polypeptide patterns of both types of membrane were studied. Many polypeptides were released from each type of membrane in all of these media. Differential effects on the polypeptide patterns of either type of membrane fraction were observed among the various buffers. In terms of minimizing loss of polypeptides, in general, NaHCO3--CacCl2 appeared to be the best buffer and Tris--HCl the worst buffer. The significance of these results for the preparation and storage of liver cell plasma membrane fractions is briefly discussed.
使用碳酸氢钠、碳酸氢钠 - 氯化钙和磷酸氢二钾 - 磷酸二氢钾缓冲液（均为pH 7.4）从大鼠肝脏中制备质膜和胆小管膜部分。使用这三种介质中的任何一种时，质膜部分的量（以每克肝脏中蛋白质的毫克数表示）都超过胆小管膜部分的量；使用碳酸氢钠 - 氯化钙可使两种类型的膜的产量显著提高。在十二烷基硫酸钠（SDS）聚丙烯酰胺凝胶电泳中，这两种膜部分呈现出复杂的多肽图谱（超过30种）。在这两种膜部分之间可观察到多肽图谱上有几个可重复的差异；特别是，可能对应于肌球蛋白重链和肌动蛋白的成分在胆小管膜部分中很突出。研究了在上述三种缓冲液和Tris - HCl（pH 7.4）中孵育对两种类型膜的多肽图谱的影响。在所有这些介质中，每种类型的膜都释放出许多多肽。在各种缓冲液中观察到对任一种膜部分的多肽图谱有不同的影响。一般而言，就尽量减少多肽损失而言，碳酸氢钠 - 氯化钙似乎是最佳缓冲液，而Tris - HCl是最差的缓冲液。简要讨论了这些结果对肝细胞质膜部分的制备和储存的意义。
