通过速率区带离心法分离的大鼠肝脏线粒体的生化异质性。

Biochemical heterogeneity of rat liver mitochondria separated by rate zonal centrifugation.

作者信息

Wilson M A, Cascarano J

出版信息

Biochem J. 1972 Aug;129(1):209-18. doi: 10.1042/bj1290209.

DOI:10.1042/bj1290209

PMID:4675005

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC1174058/

Abstract

Rat liver mitochondria were separated on the basis of their sedimentation coefficients in an iso-osmotic gradient of Ficoll-sucrose by rate zonal centrifugation. The fractions (33, each of 40ml) were collected in order of decreasing density. Fractions were analysed by spectral analysis to determine any differences in the concentrations of the cytochromes and by enzyme analyses to ascertain any differences in the activities of NADH dehydrogenase, succinate dehydrogenase and alpha-glycerophosphate dehydrogenase. 2. When plotted as% of the highest specific concentration, the contents of cytochrome a+a(3) and cytochrome c+c(1) were constant in all fractions but cytochrome b was only 65% of its maximal concentration in fraction 7 and increased with subsequent fractions. As a result, the cytochrome b/cytochrome a+a(3) ratio almost doubled between fractions 7 and 25 whereas the cytochrome c+c(1)/cytochrome a+a(3) ratio was unchanged. 3. Expression of the dehydrogenase activities as% of highest specific activity showed the following for fractions 6-26: NADH dehydrogenase activity remained fairly constant in all fractions; succinate dehydrogenase activity was 62% in fraction 6 and increased steadily to its maximum in fraction 18 and then decreased; the activity of alpha-glycerophosphate dehydrogenase was only 53% in fraction 6 and increased slowly to its peak in fractions 22 and 24. 4. These differences did not result from damaged or fragmented mitochondria or from microsomal contamination. 5. These results demonstrate that isolated liver mitochondria are biochemically heterogeneous. The importance of using a system for separating biochemically different mitochondria in studies of mitochondrial biogenesis is discussed.

摘要

通过速率区带离心法，根据大鼠肝脏线粒体在等渗的菲可-蔗糖梯度中的沉降系数对其进行分离。按密度递减顺序收集各组分（共33个，每组40毫升）。通过光谱分析确定细胞色素浓度的差异，并通过酶分析确定NADH脱氢酶、琥珀酸脱氢酶和α-甘油磷酸脱氢酶活性的差异。2. 以最高比浓度的百分比绘制时，细胞色素a+a(3)和细胞色素c+c(1)的含量在所有组分中保持恒定，但细胞色素b在第7组分中仅为其最大浓度的65%，并在后续组分中增加。结果，细胞色素b/细胞色素a+a(3)的比值在第7和第25组分之间几乎翻倍，而细胞色素c+c(1)/细胞色素a+a(3)的比值保持不变。3. 脱氢酶活性以最高比活性的百分比表示时，第6 - 26组分的情况如下：NADH脱氢酶活性在所有组分中保持相当恒定；琥珀酸脱氢酶活性在第6组分中为62%，并在第18组分中稳定增加至最大值，然后下降；α-甘油磷酸脱氢酶活性在第6组分中仅为53%，并在第22和第24组分中缓慢增加至峰值。4. 这些差异并非由受损或破碎的线粒体或微粒体污染导致。5. 这些结果表明，分离的肝脏线粒体在生化性质上具有异质性。讨论了在研究线粒体生物发生时使用一个系统来分离生化性质不同的线粒体的重要性。