Non-identical electronic characters of the internucleotidic phosphates in RNA modulate the chemical reactivity of the phosphodiester bonds†‡
Abstract
We here show that the electronic properties and the chemical reactivities of the internucleotidic phosphates in the heptameric ssRNAs are dissimilar in a sequence-specific manner because of their non-identical microenvironments, in contrast with the corresponding isosequential ssDNAs. This has been evidenced by monitoring the δH8(
) shifts upon pH-dependent ionization (pKa1) of the central 9-guaninyl (
) to the 9-guanylate ion (G−), and its electrostatic effect on each of the internucleotidic phosphate anions, as measured from the resultant δ
31P shifts (pKa2) in the isosequential heptameric ssRNAs vis-à-vis ssDNAs: [d/r(5′-Cp1Ap2Q1p3
p4Q2p5Ap6C-3′): Q1 = Q2 = A (5a/5b) or C (8a/8b), Q1 = A, Q2 = C (6a/6b), Q1 = C, Q2 = A (7a/7b)]. These oligos with single ionizable
in the centre are chosen because of the fact that the pseudoaromatic character of
can be easily modulated in a pH-dependent manner by its transformation to G− (the 2′-OH to 2-O− ionization effect is not detectable below pH 11.6 as evident from the N1-Me-
analog), thereby modulating/titrating the nature of the electrostatic interactions of
to G− with the phosphates, which therefore constitute simple models to interrogate how the variable pseudoaromatic characters of nucleobases under different sequence context (J. Am. Chem. Soc., 2004, 126, 8674–8681) can actually influence the reactivity of the internucleotide phosphates as a result of modulation of sequence context-specific electrostatic interactions. In order to better understand the impact of the electrostatic effect of the
to G− on the tunability of the electronic character of internucleotidic phosphates in the heptameric ssRNAs 5b, 6b, 7b and 8b, we have also performed their alkaline hydrolysis at pH 12.5 at 20 °C, and have identified the preferences of the cleavage sites at various phosphates, which are p2, p3 and p4 (
heptameric ssRNA sequences 5c, 7c and 8c under identical conditions in order to establish the role of the electrostatic effect of the 9-guanylate ion (and the 2′-OH to 2-O− ionization) on the internucleotidic phosphate. It turned out that the relative alkaline hydrolysis rate at those particular phosphates (p2, p3 and p4) in the N1-Me-
heptamers was reduced from 16–78% compared to those in the native counterparts [