Structural characterization of a six-nucleotide RNA hairpin loop found in Escherichia coli, r(UUAAGU)

The binding region of the Escherichia coli S2 ribosomal protein contains a conserved UUAAGU hairpin loop. The structure of the hairpin formed by the o ligomer r(GCGU4U5A6A7G8U9CGCA), which has an r(UUAAGU) hairpin loop, was de termined by NMR and molecular modeling techniques as part of a study aimed at characterizing the structure and thermodynamics of RNA hairpin loops. Th ermodynamic data obtained from melting curves for this RNA oligomer show th at it forms a hairpin in solution with the following parameters: DeltaH deg rees = -42.8 +/- 2.2 kcal/mol, DeltaS degrees = -127.6 +/- 6.5 eu, and Delt aG degrees (37) = -3.3 +/- 0.2 kcal/mol. Two-dimensional NOESY WATERGATE sp ectra show an NOE between U imino protons, which suggests that U4 and U9 fo rm a hydrogen bonded U-U pair. The U5(H2 ') proton shows NOES to both the A 6(H8) proton and the A7(H8) proton, which is consistent with formation of a "U" turn between nucleotides U5 and A6. An NOE between the A7(H2) proton a nd the U9(H4 ') proton shows the proximity of the A7 base to the U9 sugar, which is consistent with the structure determined for the six-nucleotide lo op. In addition to having a hydrogen-bonded U-U pair as the first mismatch and a U turn, the r(UUAAGU) loop has the G8 base protruding into the solven t. The solution structure of the r(UUAAGU) loop is essentially identical to the structure of an identical loop found in the crystal structure of the 3 0S ribosomal subunit where the guanine in the loop is involved in tertiary interactions with RNA bases from adjacent regions [Wimberly, B. T., Broders en, D. E., Clemons, W. M., Morgan-Warren, R. J., Carter, A. P., Vonrhein, C ., Hartsch, T., and Ramakrishnan, V. (2000) Nature 407, 327-339]. The simil arity of the solution and solid-state structures of this hairpin loop sugge sts that formation of this hairpin may facilitate folding of 16S RNA.