Estrogen Modification of Human Glutamate Dehydrogenases Is Linked to Enzyme Activation State
Estrogen Modification of Human Glutamate Dehydrogenases Is Linked to Enzyme Activation State
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DOI：
10.1074/jbc.m110.146084
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发表时间：
2010-10-08
影响因子：
4.8
通讯作者：
Plaitakis, Andreas
中科院分区：
文献类型：
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作者：
Borompokas, Nikolas;Papachatzaki, Maria-Martha;Plaitakis, Andreas
哺乳动物谷氨酸脱氢酶(GDH)是谷氨酸代谢中心的管家酶。它的活性被GTP有效地抑制(IC50=0.1-0.3微米)，并被认为是由细胞在ATP中的需要控制的。众所周知，雌激素也可以抑制哺乳动物的GDH，但浓度相对较高。因为，除了这个看家的人(H)GDH1之外，人类还通过复制事件获得了在人类皮质星形胶质细胞中表达的hGDH2亚型，因此我们在这里测试了雌激素与两种人类同工酶的相互作用。结果表明，在基线条件下，己烯雌酚对hGDH_2有较强的抑制作用(IC_(50)=0.08+/-0.01亩M)，与亲和力降低18倍的h-GDH_1相似(IC_(50)=1.67+/-0.06亩M；P&t；0.001)。同样，17β-雌二醇对hGDH2的亲和力(IC_(50)=1.53+/-0.24muM)与hGDH1的亲和力(IC_(50)=26.94+/-1.07muM；p<；0.001)接近18倍。此外，雌三醇和孕酮对hGDH2的抑制作用比hGDH1更强。结构/功能分析表明，hGDH2对雌激素的敏感性在很大程度上与R443S的进化替换有关，R443S具有较低的基础活性。雌激素对两种人类GDH的抑制与ADP诱导的激活状态呈负相关，hGDH2的这种相关性的斜率比hGDH1的更大。此外，对显示不同激活状态的hGDH1和hGDH2突变体的研究表明，雌激素对这些酶的亲和力与基础催化活性呈负相关(R=0.99；p=0.0001)。由于星形胶质细胞已知可以合成雌激素，这些激素通过与处于关闭状态的hGDH2有效地相互作用，可能有助于调节大脑中的谷氨酸代谢。
Mammalian glutamate dehydrogenase (GDH) is a housekeeping enzyme central to the metabolism of glutamate. Its activity is potently inhibited by GTP (IC50 = 0.1-0.3 mu M) and thought to be controlled by the need of the cell in ATP. Estrogens are also known to inhibit mammalian GDH, but at relatively high concentrations. Because, in addition to this housekeeping human (h) GDH1, humans have acquired via a duplication event an hGDH2 isoform expressed in human cortical astrocytes, we tested here the interaction of estrogens with the two human isoenzymes. The results showed that, under base-line conditions, diethylstilbestrol potently inhibited hGDH2 (IC50 = 0.08 +/- 0.01 mu M) and with similar to 18-fold lower affinity hGDH1 (IC50 = 1.67 +/- 0.06 mu M; p < 0.001). Similarly, 17 beta-estradiol showed a similar to 18-fold higher affinity for hGDH2 (IC50 = 1.53 +/- 0.24 mu M) than for hGDH1 (IC50 = 26.94 +/- 1.07 mu M; p < 0.001). Also, estriol and progesterone were more potent inhibitors of hGDH2 than hGDH1. Structure/function analyses revealed that the evolutionary R443S substitution, which confers low basal activity, was largely responsible for sensitivity of hGDH2 to estrogens. Inhibition of both human GDHs by estrogens was inversely related to their state of activation induced by ADP, with the slope of this correlation being steeper for hGDH2 than for hGDH1. Also, the study of hGDH1 and hGDH2 mutants displaying different states of activation revealed that the affinity of estrogen for these enzymes correlated inversely (R = 0.99; p = 0.0001) with basal catalytic activity. Because astrocytes are known to synthesize estrogens, these hormones, by interacting potently with hGDH2 in its closed state, may contribute to regulation of glutamate metabolism in brain.