Book of Abstracts: Albany 2003

category image Albany 2003
Conversation 13
Abstract Book
June 17-21 2003

A New RNA Fold: RNA Hairpins Containing Unusually Stable 2',5'-Linked r(UUCG) Loops

We have recently shown that hairpin structures of the sequence 5'-G1G2A3C4(U5U6C7G8)G9U10C11C12-3' and containing a 2',5'-linked RNA loop are of superior thermodynamic stability compared to native hairpins comprised of 3',5'-RNA loops (1,2). We have also observed, for the first time, the formation of a DNA:2',5'-RNA hybrid by linking the two strands to a 2',5'-RNA loop (1). The effect of chemical linkage on loop stability is highly surprising and enthalpic in origin. A remarkable feature of the 2',5'-r(UUCG) tetraloop is that its stability is virtually independent of the hairpin stem composition (DNA:DNA, RNA:RNA, 2',5'-RNA:2',5'-RNA, etc?). This is in sharp contrast to the native 3',5'-linked RNA tetraloop whose stability is strongly dependent on the chemistry of the stem.

To gain further insight into the molecular basis behind the unusual stability of 2',5'-linked RNA loops, we have studied the above hairpin sequence using high-resolution NMR. We show that the 2',5'-linked RNA loop adopts a new fold that is completely different from that previously observed for the native 3',5'-linked RNA loop. The 2',5'-RNA loop is stabilized by (a) U5·G8 wobble base pairing, with both nucleotide residues in the anti conformation, (b) extensive base stacking, and (c) sugar?base and sugar-sugar contacts, all of which contribute to the extra stability of this hairpin structure. The U5·G8 base pair stacks on top of the C4·G9 loop-closing base pair and thus appears as a continuation of the stem. The loop uracil (U6) base stacks on top of U5, while cytosine (C7) protrudes out into the solvent and does not participate in any of the stabilizing interactions. The different sugar pucker and intrinsic bonding interactions within the 2',5'-linked ribonucleotides help explain the unusual stability and conformational properties displayed by 2',5'-RNA tetraloops. This discovery may help in the design of more effective antisense nucleic acid agents and identifies the 2',5'-RNA loop as a novel structural motif.

Rami N. Hannoush
Aleksej Denisov
Kalle Gehring
Masad J. Damha*

Otto Maass Chemistry Building
McGill University
801 Sherbrooke St. W.
Montreal, Quebec, Canada, H3A 2K6

References and Footnotes
  1. Hannoush, R.N.; Damha, M.J. J. Am. Chem. Soc. 123, 12368 ?12374 (2001).
  2. Hannoush, R.N.; Damha, M.J. Nucleosides & Nucleotides 20, 1201-1204 (2001).
  3. (a) Cheong, C.; Varani, G.; Tinoco I. Jr. Nature 346, 680-682 (1990); (b) Varani, G.; Cheong, C.; Tinoco I. Jr. Biochemistry 30, 3280-3289 (1991); (c) Allain, F. H.-T.; Varani, G. J. Mol. Biol. 250, 333 (1995).