McKeehan W L, McKeehan K A
J Cell Physiol. 1982 Feb;110(2):142-8. doi: 10.1002/jcp.1041100206.
A sensitive isotope exchange method was developed to assess the requirements for and compartmentation of pyruvate and oxalacetate production from malate in proliferating and nonproliferating human fibroblasts. Malatedependent pyruvate production (malic enzyme activity) in the particulate fraction containing the mitochondria was dependent on either NAD+ or NADP+. The production of pyruvate from malate in the soluble, cytosolic fraction was strictly dependent on NADP+. Oxalacetate production from malate (malate dehydrogenase, EC 1.1.1.37) in both the particulate and soluble fraction was strictly dependent on NAD+. Relative to nonproliferating cells, NAD+-linked malic enzyme activity was slightly reduced and the NADP+-linked activity was unchanged in the particulate fraction of serum-stimulated, exponentially proliferating cells. However, a reduced activity of particulate malate dehydrogenase resulted in a two-fold increase in the ratio of NAD(P)+-linked malic enzyme to NAD+-linked malate dehydrogenase activity in the particulate fraction of proliferating fibroblasts. An increase in soluble NADP+-dependent malic enzyme activity and a decrease in NAD+-linked malate dehydrogenase indicated an increase in the ratio of pyruvate-producing to oxalacetate-producing malate oxidase activity in the cytosol of proliferating cells. These coordinate changes may affect the relative amount of malate that is oxidized to oxalacetate and pyruvate in proliferating cells and, therefore, the efficient utilization of glutamine as a respiratory fuel during cell proliferation.
开发了一种灵敏的同位素交换方法，以评估在增殖和非增殖的人成纤维细胞中，苹果酸产生丙酮酸和草酰乙酸的需求及区室化情况。含有线粒体的颗粒部分中依赖苹果酸的丙酮酸产生（苹果酸酶活性）依赖于NAD⁺或NADP⁺。可溶性胞质部分中苹果酸产生丙酮酸的过程则严格依赖于NADP⁺。颗粒部分和可溶性部分中苹果酸产生草酰乙酸（苹果酸脱氢酶，EC 1.1.1.37）的过程均严格依赖于NAD⁺。相对于非增殖细胞，在血清刺激的指数增殖细胞的颗粒部分中，与NAD⁺相关的苹果酸酶活性略有降低，而与NADP⁺相关的活性未发生变化。然而，颗粒部分苹果酸脱氢酶活性的降低导致增殖成纤维细胞颗粒部分中与NAD(P)⁺相关的苹果酸酶与与NAD⁺相关的苹果酸脱氢酶活性之比增加了两倍。可溶性NADP⁺依赖性苹果酸酶活性的增加以及与NAD⁺相关的苹果酸脱氢酶活性的降低表明，增殖细胞胞质中产生丙酮酸的苹果酸氧化酶活性与产生草酰乙酸的苹果酸氧化酶活性之比增加。这些协同变化可能会影响增殖细胞中苹果酸氧化为草酰乙酸和丙酮酸的相对量，从而影响细胞增殖过程中谷氨酰胺作为呼吸燃料的有效利用。
