HALOGENATED NUCLEIC ACIDS: STRUCTURE AND CONFORMATIONAL STUDIES OF 5-FLUOROCYTIDINE BY X-RAY CRYSTALLOGRAPHY AND NMR SPECTROSCOPY

Abstract

The stereochemistry and the conformational equilibria of 5-fluorocytidine (5FCyd) have been determined by X-ray crystallography and correlated with NMR spectroscopy in the solution state. Crystals of 5FCyd have unit cell dimensions a=9.854(1), b=l5.012(2), c=15.290(2)Å, α=β=γ=90° with two molecules in the asymmetric unit. Both molecules are in the anti-conformation, C3′-endo sugar pucker and a ϒ (C4′-C5′) of g⁺. The two molecules in the asymmetric unit show slight variation in their bond distances and bond angles but their overall solid state conformation is similar. The NMR results indicate the 5FCyd has an anti-conformation, a mixed sugar pucker of 36% C2′- endo and 64% C3′-endo and an exocyclic furanose conformation (ϒ) of 74%(g⁺), 19%(t) and 7%(g⁻).

Certain purine and pyrimidine analogues readily replace the natural bases in nucleic acid if they are present during replication. Halogenated nucleic acids have been known for the past thirty years when for the first time it was found that 5-iodouracil could be incorporated into the nucleic acid of E coli ¹. This provided the basis for the synthesis of 5-fluoro-nucleic acid bases and their use as a chemotherapeutic agent.

The biological properties of 5-halogenated uracil derivatives have been examined in detail. The incorporation of the halogenated base into mRNA could lead to errors in the reading frame which would alter the phenotype without permanent change in DNA genotype^2-4, also, it can substitute for uracil into mRNA and may exert a mutagenic effect⁵ or can occasionally lead to the production of altered protein^6-7. A substitution of uracil by fluorouracil in the mRNA and a preferential pairing of the 5FUra with a guanosine in the tRNA anti-codons can account for the first substitution. This is because 5FU assumes properties similar to cytosine when the base ionizes. This occurs readily for 5FU (pKa = 7.8) due to the electronegative fluorine atom at C5 instead of hydrogen. 5FCyd shows antifungal activity against Saccharomyces cerevisiae and Candida species. It is the drug of choice for systemic infections. In order to obtain a more basic understanding of the potential of such processes, the present study was undertaken as the first step using solid phase study by X-ray crystallography and its structure in solution by NMR spectroscopy⁸.