Persson T, Larsson G, Nyman P O
Department of Organic Chemistry, Lund University, Sweden.
Bioorg Med Chem. 1996 Apr;4(4):553-6. doi: 10.1016/0968-0896(96)00044-2.
The dUDP analogue, 2'-deoxyuridine 5'-(alpha,beta-imido)diphosphate (dUPNP) was synthesized. The corresponding triphosphate analogue (dUPNPP) was prepared by enzymic phosphorylation of dUPNP using the enzyme pyruvate kinase and phosphoenolpyruvate as the phosphate donor. This method was successful in phosphorylating the imidodiphosphate analogue of 2'-deoxythymidine (dTPNP) to 2'-deoxythymidine 5'-(alpha, beta-imido)triphosphate (dTPNPP), in contradiction to a previous report. The properties of dUPNPP have been tested using the enzyme dUTPase from Escherichia coli. This enzyme, having a crucial role in nucleotide metabolism, is strictly specific for its substrate (dUTP) and catalyzes the hydrolysis of the alpha, beta-bridge, resulting in dUMP and pyrophosphate. Replacement of the alpha, beta-bridging oxygen in dUTP with an imido group resulted in a nonhydrolyzable substrate analogue and a potent competitive inhibitor of dUTPase (Ki = 5 microM). The analogue prepared (dUPNPP) may be utilized in crystallographic studies of the active site of dUTPase to provide knowledge about specific interactions involved in substrate binding and as a parental compound in design of dUTPase inhibition for medical purposes.
合成了二磷酸尿苷类似物2'-脱氧尿苷5'-(α,β-亚氨基)二磷酸(dUPNP)。使用丙酮酸激酶和磷酸烯醇丙酮酸作为磷酸供体，通过酶促磷酸化dUPNP制备了相应的三磷酸类似物(dUPNPP)。与之前的报道相反，该方法成功地将2'-脱氧胸苷的亚氨基二磷酸类似物(dTPNP)磷酸化为2'-脱氧胸苷5'-(α,β-亚氨基)三磷酸(dTPNPP)。已使用来自大肠杆菌的dUTPase酶测试了dUPNPP的性质。这种在核苷酸代谢中起关键作用的酶对其底物(dUTP)具有严格的特异性，并催化α,β-桥的水解，生成dUMP和焦磷酸。用亚氨基取代dUTP中的α,β-桥连氧，得到一种不可水解的底物类似物和dUTPase的强效竞争性抑制剂(Ki = 5 μM)。所制备的类似物(dUPNPP)可用于dUTPase活性位点的晶体学研究，以提供有关底物结合中涉及的特定相互作用的知识，并作为用于医学目的的dUTPase抑制设计中的母体化合物。
