Jarmuła A, Anulewicz R, Leś A, Cyrański M K, Adamowicz L, Bretner M, Felczak K, Kulikowski T, Krygowski T M, Rode W
Nencki Institute of Experimental Biology, Polish Academy of Sciences, Warszawa, Poland.
Biochim Biophys Acta. 1998 Feb 17;1382(2):277-86. doi: 10.1016/s0167-4838(97)00169-6.
In order to understand the influence on thymidylate synthase interactions with dUMP analogues of the pyrimidine ring 2- and/or 4-thio, and 5-fluoro substitutions, X-ray diffractions by crystals of 5-fluoro-dUrd and its 2- and 4-thio, and 2,4-dithio analogues were measured, the four structures solved and refined. The following conclusions were suggested by results of comparative analyses of structural parameters (bond lengths, valence angles), followed by theoretical considerations based on calculated resonance structure distributions and aromaticity indices of the uracil, thiouracil, fluorouracil and fluorothiouracil rings. The effect of 4-thio substitution of FdUMP, altering specificity of inactivation of thymidylate synthases from various sources, is probably due to weaker proton acceptor power of the 4-thio substituent and increasing acidity (enhanced proton-donor power) of the N(3)-H moiety, resulting in an impaired fitness into the network of hydrogen bonds in the enzyme active center cleft. 2,4-Dithio substitution results in (i) impaired pyrimidine ring recognition by the enzyme active center, due to the 4-thio substituent (ii) increased pyrimidine ring aromaticity in dUMP, leading to resistance of C(6) to nucleophilic attack by the enzyme active center cysteine and (iii) altered planarity of the pyrimidine ring and deflections, with respect to the ring plane, of substituents at C(2), C(4) and C(5). 5-Fluoro substitution apparently activates the pyrimidine ring towards the interaction with thymidylate synthase by producing local strain, which results in an increased reactivity as predicted by the Walsh-Bent rule.
为了了解嘧啶环2-和/或4-硫代以及5-氟取代的dUMP类似物对胸苷酸合成酶相互作用的影响，测量了5-氟-dUrd及其2-和4-硫代以及2,4-二硫代类似物晶体的X射线衍射，解析并精修了这四种结构。通过对结构参数（键长、价键角）进行比较分析，随后基于尿嘧啶、硫代尿嘧啶、氟尿嘧啶和氟硫代尿嘧啶环的计算共振结构分布和芳香性指数进行理论考量，得出了以下结论。FdUMP的4-硫代取代改变了来自不同来源的胸苷酸合成酶的失活特异性，其作用可能是由于4-硫代取代基的质子接受能力较弱以及N(3)-H部分的酸度增加（增强的质子供体能力），导致其与酶活性中心裂隙中的氢键网络适配性受损。2,4-二硫代取代导致：(i) 由于4-硫代取代基，酶活性中心对嘧啶环的识别受损；(ii) dUMP中嘧啶环的芳香性增加，导致C(6)对酶活性中心半胱氨酸的亲核攻击具有抗性；(iii) 嘧啶环的平面性改变以及C(2)、C(4)和C(5)处取代基相对于环平面的偏转。5-氟取代显然通过产生局部应变激活了嘧啶环与胸苷酸合成酶的相互作用，这导致了如Walsh-Bent规则所预测的反应性增加。
