Laboratório de Oncobiologia Molecular (LabOMol) and Laboratório de Enzimologia e Controle do Metabolismo (LabECoM), Departamento de Fármacos, Faculdade de Farmácia, Universidade Federal do Rio de Janeiro, Rio de Janeiro, RJ 21941-590, Brazil.
Arch Biochem Biophys. 2010 May;497(1-2):62-7. doi: 10.1016/j.abb.2010.03.013. Epub 2010 Mar 25.
Clotrimazole (CTZ) has been proposed as a potential anti-neoplastic agent, which inhibits glucose metabolism. The present work aimed to evaluate the effects of CTZ on the kinetic mechanism of 6-phosphofructo-1-kinase (PFK). We show that CTZ promotes a dose-dependent inhibition of PFK, presenting a K(i) of 28 +/- 2 microM. Inhibition occurs through the dissociation of the enzyme tetramers, as demonstrated through fluorescence spectroscopy and gel filtration chromatography. Moreover, the affinities of the enzyme for ATP and fructose-6-phosphate are reduced 50% and 30%, respectively. Furthermore, the affinity of PFK for ATP at the inhibitory site becomes 2-fold higher. Altogether, the results presented here suggest that PFK inhibition by CTZ involves a decrease in the affinity of PFK for its substrates at the catalytic site with the concomitant potentiation of the inhibitory properties of ATP.
克霉唑（CTZ）已被提议作为一种潜在的抗肿瘤药物，它能抑制葡萄糖代谢。本工作旨在评估 CTZ 对 6-磷酸果糖-1-激酶（PFK）动力学机制的影响。我们表明 CTZ 能促进 PFK 的剂量依赖性抑制，其 K(i)值为 28 ± 2 μM。抑制作用是通过酶四聚体的解离来实现的，这一点通过荧光光谱和凝胶过滤色谱得到了证明。此外，酶对 ATP 和果糖-6-磷酸的亲和力分别降低了 50%和 30%。此外，PFK 在抑制部位对 ATP 的亲和力增加了 2 倍。总之，这里提出的结果表明，CTZ 对 PFK 的抑制作用涉及到降低 PFK 对其在催化部位的底物的亲和力，同时增强 ATP 的抑制特性。
