Chang G G, Wang J K, Huang T M, Lee H J, Chou W Y, Meng C L
Department of Biochemistry, National Defense Medical Center, Taipei, Taiwan, Republic of China.
Eur J Biochem. 1991 Dec 5;202(2):681-8. doi: 10.1111/j.1432-1033.1991.tb16423.x.
Cytosolic NADP(+)-dependent malic enzyme from a cultured human breast cancer cell line was purified to near homogeneity by two highly efficient chromatography systems: Pharmacia-LKB Q-Sepharose anion-exchange chromatography and adenosine-2',5'-bisphosphate-agarose affinity chromatography. The overall yield was 27%. The enzyme is presumably a tetramer composed of four probably identical subunits of Mr 65,000, which is similar to the enzyme from other sources. The pI and optimum reaction pH values for the tumor malic enzyme are 5.5 and 7.2, respectively. At pH 6.9, most of the enzyme exists as monomers. Activation energy for the enzyme-catalyzed oxidative-decarboxylation reaction is 57.4 kJ/mol. The enzyme is strictly NADP+ dependent, as NAD+ cannot support the oxidative-decarboxylation reaction. ATP at low concentration inhibits the enzyme activity. Fumarate at concentrations up to 5 mM does not affect the enzymatic reaction rate. Therefore the tumor cytosolic malic enzyme, unlike the mitochondrial malic enzyme, is not an allosteric regulatory enzyme.
通过两个高效的色谱系统，即Pharmacia-LKB Q-琼脂糖阴离子交换色谱和2',5'-二磷酸腺苷-琼脂糖亲和色谱，从培养的人乳腺癌细胞系中纯化出胞质NADP(+)-依赖性苹果酸酶，使其接近均一。总产率为27%。该酶可能是由四个Mr 65,000的可能相同的亚基组成的四聚体，这与其他来源的酶相似。肿瘤苹果酸酶的pI和最佳反应pH值分别为5.5和7.2。在pH 6.9时，大多数酶以单体形式存在。酶催化的氧化脱羧反应的活化能为57.4 kJ/mol。该酶严格依赖NADP+，因为NAD+不能支持氧化脱羧反应。低浓度的ATP抑制酶活性。浓度高达5 mM的富马酸盐不影响酶促反应速率。因此，肿瘤胞质苹果酸酶与线粒体苹果酸酶不同，不是变构调节酶。
