Fig 2.

Reactivity of RNA and RNA analogs. (A) Relative electrophoretic mobilities of 2′-amino and 2′-hydroxyl nucleotides visualized by autoradiography of 5′-³²P-end-labeled molecules (Left) or UV shadowing (Right). (B) 2′-Amine acylation reactions for the reference dinucleotide pU^2′-NH₂pU (1), the all-ribose control pU^2′-OHpU (2), and the phosphotriester (8), resolved by gel electrophoresis. Filled arrows indicate 2′-acylated product, and open arrows designate background modification for 2 and a degradative by-product for 8. (C) Reaction of a succinimidyl ester with RNA is selective for a 2′-amine substitution and is enhanced in flexible structures. A 20-mer single-stranded oligonucleotide (15) containing a 2′-amine substitution at position 10 (open circles, k_obs = 0.077 min⁻¹) reacts ≈5-fold and 20-fold faster than if constrained in a mismatch-containing or duplex helix (open diamonds and squares, respectively). (D) Reaction of cAMP^2′-NH₂ (9) with a succinimidyl ester is undetectable, as judged by UV shadowing. In contrast, U^2′-NH₂pU (3) reacts to form the 2′-amide product (arrow) under the same conditions. (E) Reaction of xyl-T^2′-NH₂ (12) and U^2′-NH₂ (11) monitored by formation of an anionic fluorescein adduct by using gel electrophoresis. U and − indicate reactions performed in the presence of the all-ribose uridine nucleoside (U^2′-OH) and in the absence of nucleoside, respectively. Arrows indicate two product bands for U^2′-NH₂, corresponding to regioisomers of the fluorescein-conjugated succinimidyl ester; asterisks indicate background bands observed in the no nucleoside (−) control. The fluorescently labeled xyl-T^2′-NH₂ product is undetectable.