Mackinnon W B, Mountford C E
Ludwig Institute for Cancer Research, University of Sydney, New South Wales, Australia.
Anal Biochem. 1988 Dec;175(2):386-9. doi: 10.1016/0003-2697(88)90561-1.
The purity of isolated plasma membranes is routinely judged by the activity of enzymes present both in this membrane and other locations in the cell. However, since enzyme inhibition and/or stimulation often occurs following disruption of the cell, the question as to which enzyme(s) provides a reliable marker of membrane purity should be considered. We have devised a simple method with which to address this problem. Inhibition or stimulation of plasma membrane marker enzymes can be rapidly assessed in cell homogenates and subfractions by mixing both samples, with known enzyme activity, and observing any deviation from the expected combined activity. Should the activity remain constant that enzyme can be used to gauge the purity of the plasma membrane preparation. Of the four putative plasma membrane marker enzymes examined only one, gamma-glutamyltranspeptidase appeared to give a reliable purity measurement in the cell system studied.
分离得到的质膜的纯度通常通过存在于该膜以及细胞其他位置的酶的活性来判断。然而，由于细胞破裂后常常会发生酶抑制和/或刺激作用，因此应该考虑哪种酶能够提供可靠的膜纯度标记这一问题。我们设计了一种简单的方法来解决这个问题。通过将具有已知酶活性的细胞匀浆和亚组分混合，并观察与预期的综合活性的任何偏差，可以快速评估质膜标记酶的抑制或刺激情况。如果活性保持恒定，则该酶可用于衡量质膜制备物的纯度。在所研究的四种假定的质膜标记酶中，只有γ-谷氨酰转肽酶似乎在所研究的细胞系统中给出了可靠的纯度测量结果。
