Graham James W A, Williams Thomas C R, Morgan Megan, Fernie Alisdair R, Ratcliffe R George, Sweetlove Lee J
Department of Plant Sciences, University of Oxford, Oxford, OX1 3RB, United Kingdom.
Plant Cell. 2007 Nov;19(11):3723-38. doi: 10.1105/tpc.107.053371. Epub 2007 Nov 2.
In Arabidopsis thaliana, enzymes of glycolysis are present on the surface of mitochondria and free in the cytosol. The functional significance of this dual localization has now been established by demonstrating that the extent of mitochondrial association is dependent on respiration rate in both Arabidopsis cells and potato (Solanum tuberosum) tubers. Thus, inhibition of respiration with KCN led to a proportional decrease in the degree of association, whereas stimulation of respiration by uncoupling, tissue ageing, or overexpression of invertase led to increased mitochondrial association. In all treatments, the total activity of the glycolytic enzymes in the cell was unaltered, indicating that the existing pools of each enzyme repartitioned between the cytosol and the mitochondria. Isotope dilution experiments on isolated mitochondria, using (13)C nuclear magnetic resonance spectroscopy to monitor the impact of unlabeled glycolytic intermediates on the production of downstream intermediates derived from (13)C-labeled precursors, provided direct evidence for the occurrence of variable levels of substrate channeling. Pull-down experiments suggest that interaction with the outer mitochondrial membrane protein, VDAC, anchors glycolytic enzymes to the mitochondrial surface. It appears that glycolytic enzymes associate dynamically with mitochondria to support respiration and that substrate channeling restricts the use of intermediates by competing metabolic pathways.
在拟南芥中，糖酵解酶存在于线粒体表面和胞质溶胶中。通过证明线粒体结合程度取决于拟南芥细胞和马铃薯块茎中的呼吸速率，现已确定了这种双重定位的功能意义。因此，用氰化钾抑制呼吸会导致结合程度成比例下降，而通过解偶联、组织老化或转化酶过表达刺激呼吸会导致线粒体结合增加。在所有处理中，细胞中糖酵解酶的总活性未改变，表明每种酶的现有库在胞质溶胶和线粒体之间重新分配。使用（13）C核磁共振光谱监测未标记的糖酵解中间体对源自（13）C标记前体的下游中间体产生的影响，对分离的线粒体进行同位素稀释实验，为底物通道化水平变化的发生提供了直接证据。下拉实验表明，与线粒体外膜蛋白VDAC的相互作用将糖酵解酶锚定在线粒体表面。糖酵解酶似乎与线粒体动态结合以支持呼吸，并且底物通道化通过竞争代谢途径限制了中间体的使用。
