Halogenated nucleic acids: effects of 5-fluorouracil on the conformation and properties of a polyribonucleotide and its constituents

The conformational properties of 5-fluorouracil derivatives are compared to uracil derivatives. FUrd, 5′-FUMP, and poly(FU) are studied as a function of pH and temperature by ¹⁹F- and ¹H-nmr spectroscopy, and the corresponding uracil derivatives by ¹H-nmr spectroscopy. FUrd exhibits no significant conformational changes with solution pH (5–10). In contrast, at low pH (6–7) 5′-FUMP and 5′-UMP show similar conformational features, while at high pH (9) 5′-FUMP shows significant conformational alterations. Also, poly(U) and poly(FU) are conformationally similar at low pH, but increasing pH induces changes in poly(FU). These changes are observed in the backbone γ(C4′-C5′)], furanose, and furanose-base conformations. The apparent pK_a of N3-H ionization of the FUra base is determined by ¹H- and ¹⁹F-nmr to range from 7.5 to 8.2 FUrd < 5′-FUMP < 5′-FUDP < poly(FU)]. These observations are interpreted as a result of electrostatic interactions generated between the ionized phosphate group and the negatively charged base moiety as the pH is raised. The interaction properties of poly(FU) with ApA are studied by ¹H- and ¹⁹F-nmr spectroscopy, and these properties compared to those published for poly(U). Poly(FU) forms a complex with ApA inducing upfield ¹H-shifts in both components, and downfield ¹⁹F- shifts in poly(FU). The base stoichiometry of the complex for poly(U)·ApA is 2U:1A at various U/A ratios. In contrast, the base stoichiometry of the poly(FU)·ApA complex appears to be dependent on the FU/A ratio. At high FU/A ratio, the complex is 2FU:1A, and as the FU/A ratio approaches unity the complex becomes 1FU:1A.