Significance of Mitochondrial Binding and Release of Hexokinase in Metabolic Regulation

研究了己糖激酶与线粒体结合的意义，特别是在高酶解活性的组织和细胞中，并特别参考了葡萄糖代谢的调节机制。1)细胞内阳离子条件，特别是钾离子浓度和镁离子浓度对己糖激酶的细胞内分布有显著影响。2)与线粒体的结合不仅促进了己糖激酶的作用，而且使己糖激酶稳定。3)与研究最广泛的脑内己糖激酶分布相比，我们也几乎首次在培养的神经母细胞瘤细胞中检测了己糖激酶的细胞内分布。在神经母细胞瘤细胞中发现约30%的己糖激酶活性以线粒体结合形式存在。我们还发现，己糖激酶I在细胞中占主导地位，就像在大脑中一样。4)在本研究中，己糖激酶I对线粒体的亲和力比己糖激酶II高得多，并且与细胞外葡萄糖浓度的变化无关，结合的己糖激酶几乎是恒定的。己糖激酶I的这些特性可能对大脑中葡萄糖代谢的稳态稳定很重要。5)正常大鼠肝脏和再生肝中线粒体几乎没有己糖激酶活性。然而，在一些大鼠腹水肝癌细胞的线粒体部分中发现了一定程度的活性，表明细胞内己糖激酶分布的变化与癌变有关。6)研究了己糖激酶的翻译后加工过程，假设该加工过程消除了结合结构域，导致肝脏中线粒体结合的己糖激酶缺失。在己糖激酶分子的n端存在富含疏水氨基酸的结合域，因为轻微的胰凝乳蛋白酶处理会导致线粒体结合能力的丧失，而催化活性和分子量几乎没有变化。在肝脏中发现了类似的活性，但在大脑中没有，活性集中在肝脏的溶酶体部分。从抑制剂和激活剂的研究中，发现硫醇蛋白酶负责处理。加工活性在一定程度上位于溶酶体的外表面，并讨论了其可能的加工机制。少

Significance of the binding of hexokinase to mitochondria, especially in tissues and cells with high blycolytic activity, was examined with special reference to regulatory mechanism for glucose metabolism.1) Intracellular conditions for cations, especially the concentration of potassium ion in addition to that of magnesium ion, were significantly involved in the intracellular distribution of hexokinase.2) The binding to mitochondria brought about not only the facilitation for the action but also the stabilization of hexokinase.3) Intracellular distridution of hexokinase was also examined for cultured neuroblastoma cells for almost the first time, in reference to that in the brain which has been studied most extenvsively. About 30% of hexokinase activity was found as mitochondria-bound-form in-the neuroblastoma cells. It was also found that hexokinase I was predominant in the cells as was in the brain.4) Throughout the present study, hexokinase I showed much higher affinity to mitochond … More ria than hexokinase II, and the binding hexokinase was almost constant irrespective of the change in extra-cellular concentration of gluose. These properties of hexokinase I may be important for homeo-stasis of glucose metabolism in the brain.5) Almost no hexokinase activity was bound to mitochondria in normal rat liver as well as in the regenerating liver. However, the activity was found to some extent in the mitochondria fraction of some strains of rat ascites hepatoma cells, indicating the change in the intracellular distribution of hexokinase in relation to carcinogenesis.6) Posttranslational processing of hexokinase was investigated on an assumption that the elimination of the binding domain by the processing was responsible for the absence of mitochondria-bound hexokinase in the liver. The binding domain rich in hydrophobic amino acids has been postulated in the n-terminus of hexokinase molecule, since mild chymotrypsin treatment causes loss of the mitochondria-binding ability with little changes in the catalytic activity and molecular weight. Similar activity was found in the liver but not in the brain, and the activity was concentrated in the lysosomal fraction of the liver. From inhibitor and activator studies, thiol protease was found to be responsible for the processing. The processing activity was located in the outer surface of lysosomes to some extent, and possible mechanism for the processing was discussed. Less